| 1. |
Fahmida Habib Nabila, Rashedul Islam, Islam Md Shimul, Muhammad Moniruzzaman, Rie Wakabayashi, Noriho Kamiya, Masahiro Goto, Ionic liquid-mediated ethosome for transdermal delivery of insulin., Chemical communications (Cambridge, England), 10.1039/d3cc06130b, 2024.03, Herein, we report ethosome (ET) formulations composed of a safe amount of 1,2-dimyristoyl-sn-glycero-3-phosphatidylcholine (DMPC)-based ionic liquid with various concentrations of ethanol as a carrier for the transdermal delivery of a high molecular weight drug, insulin. The Insulin-loaded ET vesicles exhibited long-term stability compared to conventional DMPC ETs, showing significantly higher drug encapsulation efficiency and increased skin permeation ability.. |
| 2. |
Ryutaro Ariyoshi, Takashi Matsuzaki, Ryo Sato, Kosuke Minamihata, Kounosuke Hayashi, Taisei Koga, Kensei Orita, Riko Nishioka, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya, Engineering the Propeptide of Microbial Transglutaminase Zymogen: Enabling Substrate-Dependent Activation for Bioconjugation Applications., Bioconjugate chemistry, 10.1021/acs.bioconjchem.3c00544, 2024.02, Microbial transglutaminase (MTG) from Streptomyces mobaraensis is a powerful biocatalytic glue for site-specific cross-linking of a range of biomolecules and synthetic molecules that have an MTG-reactive moiety. The preparation of active recombinant MTG requires post-translational proteolytic digestion of a propeptide that functions as an intramolecular chaperone to assist the correct folding of the MTG zymogen (MTGz) in the biosynthesis. Herein, we report engineered active zymogen of MTG (EzMTG) that is expressed in soluble form in the host Escherichia coli cytosol and exhibits cross-linking activity without limited proteolysis of the propeptide. We found that the saturation mutagenesis of residues K10 or Y12 in the propeptide domain generated several active MTGz mutants. In particular, the K10D/Y12G mutant exhibited catalytic activity comparable to that of mature MTG. However, the expression level was low, possibly because of decreased chaperone activity and/or the promiscuous substrate specificity of MTG, which is potentially harmful to the host cells. The K10R/Y12A mutant exhibited specific substrate-dependent reactivity toward peptidyl substrates. Quantitative analysis of the binding affinity of the mutated propeptides to the active site of MTG suggested an inverse relationship between the binding affinity and the catalytic activity of EzMTG. Our proof-of-concept study provides insights into the design of a new biocatalyst using the MTGz as a scaffold and a potential route to high-throughput screening of EzMTG mutants for bioconjugation applications.. |
| 3. |
Diah Anggraini Wulandari, Kyosuke Tsuru, Kosuke Minamihata, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya, A Functional Hydrogel Bead-Based High-Throughput Screening System for Mammalian Cells with Enhanced Secretion of Therapeutic Antibodies., ACS Biomaterials Science & Engineering, 10.1021/acsbiomaterials.3c01386, 10, 1, 628-636, 2024.01, Droplet-based high-throughput screening systems are an emerging technology that provides a quick test to screen millions of cells with distinctive characteristics. Biopharmaceuticals, specifically therapeutic proteins, are produced by culturing cells that secrete heterologous recombinant proteins with different populations and expression levels; therefore, a technology to discriminate cells that produce more target proteins is needed. Here, we present a droplet-based microfluidic strategy for encapsulating, screening, and selecting target cells with redox-responsive hydrogel beads (HBs). As a proof-of-concept study, we demonstrate the enrichment of hybridoma cells with enhanced capability of antibody secretion using horseradish peroxidase (HRP)-catalyzed hydrogelation of tetra-thiolate poly(ethylene glycol); hybridoma cells were encapsulated in disulfide-bonded HBs. Recombinant protein G or protein M with a C-terminal cysteine residue was installed in the HBs via disulfide bonding to capture antibodies secreted from the cells. HBs were fluorescently stained by adding the protein L-HRP conjugate using a tyramide signal amplification system. HBs were then separated by fluorescence-activated droplet sorting and degraded by reducing the disulfide bonds to recover the target cells. Finally, we succeeded in the selection of hybridoma cells with enhanced antibody secretion, indicating the potential of this system in the therapeutic protein production.. |
| 4. |
Ryutaro Ariyoshi, Takashi Matsuzaki, Ryo Sato, Kosuke Minamihata, Kounosuke Hayashi, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya, Engineered Active Zymogen of Microbial Transglutaminase, bioRxiv preprint doi: https://doi.org/10.1101/2023.10.09.561484, https://doi.org/10.1101/2023.10.09.561484, 2023.09. |
| 5. |
Yoshiro Tahara, Riko Mizuno, Tomoki Nishimura, Sada atsu Mukai, Rie Wakabayashi, Noriho Kamiya, Kazunari Akiyoshi, Masahiro Goto, A solid-in-oil-in-water emulsion: An adjuvant-based immune-carrier enhances vaccine effect, Biomaterials, 10.1016/j.biomaterials.2022.121385, 282, 2022.03. |
| 6. |
Kato Y, Nishiyama K, Lee JM, Ibuki Y, Imai Y, Noda T, Kamiya N, Kusakabe T, Kanda Y, Nishida M., TRPC3-Nox2 Protein Complex Formation Increases the Risk of SARS-CoV-2 Spike Protein-Induced Cardiomyocyte Dysfunction through ACE2 Upregulation., International Journal of Molecular Sciences, 10.3390/ijms24010102, 24, 1, 2022.12. |
| 7. |
Pugoh Santoso, Takuya Komada, Yugo Ishimine, Hiromasa Taniguchi, Kosuke Minamihata, Masahiro Goto, Toki Taira, Noriho Kamiya , Preparation of amphotericin B-loaded hybrid liposomes and the integration of chitin-binding proteins for enhanced antifungal activity, Journal of Bioscience and Bioengineering, 10.1016/j.jbiosc.2022.06.005, 2022.09. |
| 8. |
Hiromasa Taniguchi, Yugo Ishimime, Kosuke Minamihata, Pugoh Santoso, Takuya Komada, Hendra Saputra, Kazuki Uchida, Masahiro Goto, Toki Taira, Noriho Kamiya , Liposomal Amphotericin B Formulation Displaying Lipid-Modified Chitin-Binding Domains with Enhanced Antifungal Activity, Molecular Pharmaceutics, 10.1021/acs.molpharmaceut.2c00388, 2022.09. |
| 9. |
Rie Wakabayashi, Rino Imatani, Mutsuhiro Katsuya, Yuji Higuchi, Hiroshi Noguchi, Noriho Kamiya, Masahiro Goto , Hydrophobic immiscibility controls self-sorting or co-assembly of peptide amphiphiles, Chemical Communications, 10.1039/d1cc05560g, 58, 4, 585-588, 2022.01. |
| 10. |
Pugoh Santoso, Kosuke Minamihata, Yugo Ishimine, Hiromasa Taniguchi, Takuya Komada, Ryo Sato, Masahiro Goto, Tomoya Takashima, Toki Taira, Noriho Kamiya , Enhancement of the Antifungal Activity of Chitinase by Palmitoylation and the Synergy of Palmitoylated Chitinase with Amphotericin B, ACS Infectious Diseases, https://doi.org/10.1021/acsinfecdis.2c00052, 8, 5, 1051-1061, 2022.04. |
| 11. |
Kazuki Uchida, Hiroki Obayashi, Kosuke Minamihata, Rie Wakabayashi, Masahiro Goto, Naofumi Shimokawa, Masahiro Takagi, Noriho Kamiya, Artificial Palmitoylation of Proteins Controls the Lipid Domain-Selective Anchoring on Biomembranes and the Raft-Dependent Cellular Internalization, Langmuir, 10.1021/acs.langmuir.2c01205, 2022.08. |
| 12. |
Hori, Katsutoshi; Yoshimoto, Shogo; Yoshino, Tomoko; Zako, Tamotsu; Hirao, Gen; Fujita, Satoshi; Nakamura, Chikashi; Yamagishi, Ayana; Kamiya, Noriho, Recent advances in research on biointerfaces: From cell surfaces to artificial interfaces, J. Biosci. Bioeng., 10.1016/j.jbiosc.2021.12.004, 133, 3, 195-207, 2022.03. |
| 13. |
Ryosuke Kaneko, Tsuyoshi Oda, Ryosuke Yoshida, Chuya Tateishi, Kenta Tanito, Teruki Nii, Akihiro Kishimura, Noriho Kamiya, Takeshi Mori, Yoshiki Katayama, alpha-L-Arabinofuranosidase as an Orthogonal Enzyme for Human Cells, CHEMISTRY LETTERS, 10.1246/cl.210231, 50, 8, 1493-1495, 2021.08, Here we proposed a-L-arabinofuranosidase from Clostridium thermocellum as an enzyme for signal amplification in bioanalysis whose activity does not exist in human cells. Combination of this enzyme and beta-galactosidase enabled simultaneous detection of two different antigens on live cells.. |
| 14. |
K. Minamihata, Y. Tanaka, P. Santoso, M. Goto, D. Kozome, T. Taira, N. Kamiya, Orthogonal Enzymatic Conjugation Reactions Create Chitin Binding Domain Grafted Chitinase Polymers with Enhanced Antifungal Activity, Bioconjugate Chem., 10.1021/acs.bioconjchem.1c00235, 32, 8, 1688-1698, 2021.08. |
| 15. |
R. Wakabayashi, A. Higuchi, H. Obayashi, M. Goto, N. Kamiya, pH-Responsive Self-Assembly of Designer Aromatic Peptide Amphiphiles and Enzymatic Post-Modification of Assembled Structures, Int. J. Mol. Sci., 10.3390/ijms22073459, 22, 7, 2021.04. |
| 16. |
Y. Kato, S. Yamada, K. Nishiyama, A. Satsuka, S. Re, D. Tomokiyo, J.-M. Lee, T. Tanaka, A. Nishimura, K. Yonemitsu, H. Asakura, Y. Ibuki, Y. Imai, N. Kamiya, K. Mizuguchi, T. Kusakabe, Y. Kanda*, M. Nishida*, Clomipramine suppresses ACE2-mediated SARS-CoV-2 entry, bioRxiv preprint, https://doi.org/10.1101/2021.03.13.435221, 2021.03. |
| 17. |
Yuya Hirakawa, Hiroshi Ueda, Yusuke Takata, Kosuke Minamihata, Rie Wakabayashi, Noriho Kamiya, Masahiro Goto, Co-amorphous formation of piroxicam-citric acid to generate supersaturation and improve skin permeation., European journal of pharmaceutical sciences : official journal of the European Federation for Pharmaceutical Sciences, 10.1016/j.ejps.2020.105667, 158, 105667-105667, 2021.03, The objective of this study was to prepare a co-amorphous formulation of piroxicam (PIR), a non-steroidal anti-inflammatory drug, and citric acid (CA), and evaluate its skin permeation ability. A spray-drying method was employed to prepare the co-amorphous formulation and its physical properties were characterized. X-ray powder diffraction and thermal analysis confirmed a homogeneous amorphous state, and the infrared spectra revealed intermolecular interactions between PIR and CA, suggesting formation of a co-amorphous formulation of PIR and CA. The PIR-CA co-amorphous formulation exhibited no crystallization for 60 days at 4/25/40°C with silica gel. The PIR-CA co-amorphous formulation increased the solubility of PIR in polyethylene glycol 400 compared with that of the pure drug, and physical mixture (PM) of PIR and CA, confirming a supersaturated state in the formulation. The PIR-CA co-amorphous formulation demonstrated higher skin permeation than PIR alone or PM of PIR and CA, and the flux value was consistent with the degree of saturation. Thus, the increase in the skin permeation of PIR from the PIR-CA co-amorphous formulation directly depended on the increased thermodynamic activity by supersaturation in the absence of interactions between the drug and co-former in the vehicle.. |
| 18. |
Rie Wakabayashi, Ayato Higuchi, Hiroki Obayashi, Masahiro Goto, Noriho Kamiya, pH-Responsive Self-Assembly of Designer Aromatic Peptide Amphiphiles and Enzymatic Post-Modification of Assembled Structures., International journal of molecular sciences, 10.3390/ijms22073459, 22, 7, 2021.03, Supramolecular fibrous materials in biological systems play important structural and functional roles, and therefore, there is a growing interest in synthetic materials that mimic such fibrils, especially those bearing enzymatic reactivity. In this study, we investigated the self-assembly and enzymatic post-modification of short aromatic peptide amphiphiles (PAs), Fmoc-LnQG (n = 2 or 3), which contain an LQG recognition unit for microbial transglutaminase (MTG). These aromatic PAs self-assemble into fibrous structures via π-π stacking interactions between the Fmoc groups and hydrogen bonds between the peptides. The intermolecular interactions and morphologies of the assemblies were influenced by the solution pH because of the change in the ionization states of the C-terminal carboxy group of the peptides. Moreover, MTG-catalyzed post-modification of a small fluorescent molecule bearing an amine group also showed pH dependency, where the enzymatic reaction rate was increased at higher pH, which may be because of the higher nucleophilicity of the amine group and the electrostatic interaction between MTG and the self-assembled Fmoc-LnQG. Finally, the accumulation of the fluorescent molecule on these assembled materials was directly observed by confocal fluorescence images. Our study provides a method to accumulate functional molecules on supramolecular structures enzymatically with the morphology control.. |
| 19. |
Dani PERMANA, Kosuke MINAMIHATA, Masahiro GOTO, Noriho KAMIYA, Strategies for Making Multimeric and Polymeric Bifunctional Protein Conjugates and Their Applications as Bioanalytical Tools, Anal. Sci., https://doi.org/10.2116/analsci.20SCR07, 37, 3, 425-437, 2021.03. |
| 20. |
Ryo Sato, Kosuke Minamihata, Ryutaro Ariyoshi, Hiromasa Taniguchi, Noriho Kamiya, Recombinant production of active microbial transglutaminase in E. coli by using self-cleavable zymogen with mutated propeptide, Protein Expression and Purification, 10.1016/j.pep.2020.105730, 2020.12, Microbial transglutaminase from Streptomyces mobaraensis (MTG) has been widely used in food industry and also in research and medical applications, since it can site-specifically modify proteins by the cross-linking reaction of glutamine residue and the primary amino group. The recombinant expression system of MTG in E. coli provides better accessibility for the researchers and thus can promote further utilization of MTG. Herein, we report production of active and soluble MTG in E. coli by using a chimeric protein of tobacco etch virus (TEV) protease and MTG zymogen. A chimera of TEV protease and MTG zymogen with native propeptide resulted in active MTG contaminated with cleaved propeptide due to the strong interaction between the propeptide and catalytic domain of MTG. Introduction of mutations of K9R and Y11A to the propeptide facilitated dissociation of the cleaved propeptide from the catalytic domain of MTG and active MTG without any contamination of the propeptide was obtained. The specific activity of the active MTG was 22.7 ± 2.6 U/mg. The successful expression and purification of active MTG by using the chimera protein of TEV protease and MTG zymogen with mutations in the propeptide can advance the use of MTG and the researches using MTG mediated cross-linking reactions.. |
| 21. |
Ryo Sato, Kosuke Minamihata, Ryutaro Ariyoshi, Hiromasa Taniguchi, Noriho Kamiya, Recombinant production of active microbial transglutaminase in E. coli by using self-cleavable zymogen with mutated propeptide., Protein expression and purification, 10.1016/j.pep.2020.105730, 176, 105730-105730, 2020.12, Microbial transglutaminase from Streptomyces mobaraensis (MTG) has been widely used in food industry and also in research and medical applications, since it can site-specifically modify proteins by the cross-linking reaction of glutamine residue and the primary amino group. The recombinant expression system of MTG in E. coli provides better accessibility for the researchers and thus can promote further utilization of MTG. Herein, we report production of active and soluble MTG in E. coli by using a chimeric protein of tobacco etch virus (TEV) protease and MTG zymogen. A chimera of TEV protease and MTG zymogen with native propeptide resulted in active MTG contaminated with cleaved propeptide due to the strong interaction between the propeptide and catalytic domain of MTG. Introduction of mutations of K9R and Y11A to the propeptide facilitated dissociation of the cleaved propeptide from the catalytic domain of MTG and active MTG without any contamination of the propeptide was obtained. The specific activity of the active MTG was 22.7 ± 2.6 U/mg. The successful expression and purification of active MTG by using the chimera protein of TEV protease and MTG zymogen with mutations in the propeptide can advance the use of MTG and the researches using MTG mediated cross-linking reactions.. |
| 22. |
Shihab Uddin, Md Raihan Chowdhury, Rie Wakabayashi, Noriho Kamiya, Muhammad Moniruzzaman, Masahiro Goto, Lipid based biocompatible ionic liquids: synthesis, characterization and biocompatibility evaluation., Chemical communications (Cambridge, England), 10.1039/d0cc04491a, 56, 89, 13756-13759, 2020.11, We report a new series of lipid-based biocompatible ionic liquids (LBILs) consisting of the long-chain phosphonium compound 1,2-dimyristoyl-sn-glycero-3-ethyl-phosphatidylcholine as the cation and the long-chain fatty acids stearic acid, oleic acid, or linoleic acid as anions. These materials were found to be completely miscible with many polar and nonpolar organic solvents as well as dispersible in water. These LBILs also exhibited excellent biocompatibility with an artificial three-dimensional human epidermis model.. |
| 23. |
Wahyu Ramadhan, Yuki Ohama, Kosuke Minamihata, Kousuke Moriyama, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya, Redox-responsive functionalized hydrogel marble for the generation of cellular spheroids, Journal of Bioscience and Bioengineering, 10.1016/j.jbiosc.2020.05.010, 130, 4, 416-423, 2020.10, Liquid marbles (LMs) have recently shown a great promise as microbioreactors to construct self-supported aqueous compartments for chemical and biological reactions. However, the evaporation of the inner aqueous liquid core has limited their application, especially in studying cellular functions. Hydrogels are promising scaffolds that provide a spatial environment suitable for three-dimensional cell culture. Here, we describe the fabrication of redox-responsive hydrogel marbles (HMs) as a three-dimensional cell culture platform. The HMs are prepared by introducing an aqueous mixture of a tetra-thiolated polyethylene glycol (PEG) derivative, thiolated gelatin (Gela-SH), horseradish peroxidase, a small phenolic compound, and human hepatocellular carcinoma cells (HepG2) to the inner aqueous phase of LMs. Eventually, HepG2 cells are encapsulated in the HMs then immersed in culture media, where they proliferate and form cellular spheroids. Experimental results show that the Gela-SH concentration strongly influences the physicochemical and microstructure properties of the HMs. After 6 days in culture, the spheroids were recovered from the HMs by degrading the scaffold, and examination showed that they had reached up to about 180 μm in diameter depending on the Gela-SH concentration, compared with 60 μm in conventional HMs without Gela-SH. After long-term culture (over 12 days), the liver-specific functions (secretion of albumin and urea) and DNA contents of the spheroids cultured in the HMs were elevated compared with those cultured in LMs. These results suggest that the developed HMs can be useful in designing a variety of microbioreactors for tissue engineering applications.. |
| 24. |
K. Minamihata*, Y. Hamada, G. Kagawa, W. Ramadhan, A. Higuchi, K. Moriyama, R. Wakabayashi, M. Goto, N. Kamiya*, Dual-Functionalizable Streptavidin–SpyCatcher-Fused Protein–Polymer Hydrogels as Scaffolds for Cell Culture, ACS Appl. Bio Mater., https://doi.org/10.1021/acsabm.0c00940, 3, 7734-7742, 2020.10. |
| 25. |
Yoshiro Tahara, Kaho Morita, Rie Wakabayashi, Noriho Kamiya, Masahiro Goto, Biocompatible Ionic Liquid Enhances Transdermal Antigen Peptide Delivery and Preventive Vaccination Effect., Molecular pharmaceutics, 10.1021/acs.molpharmaceut.0c00598, 17, 10, 3845-3856, 2020.10, Ionic liquids (ILs) attract significant attention as novel solvents for drug delivery systems because of their ability to solubilize poorly soluble drugs and tune the physiological properties of active pharmaceutical ingredients. For the next generation of IL-based drug delivery systems, biocompatibility is a high priority. In the current study, choline-fatty acids ([Cho][FA]) were used as a biocompatible IL to mediate the dissolution of a water-soluble antigen peptide in an oil-based skin penetration enhancer. Among the candidate fatty acids (C8, C10, C12, C14, C16, C18:0, and C18:1), C18:1 was selected because of its low cytotoxicity and mediation of skin permeability for an antigen peptide. Using IL[Cho][C18:1] and an oil-based penetration enhancer, the flux of transdermal delivery of the peptide increased 28-fold compared with delivery using an aqueous vehicle. Furthermore, the IL-mediated transcutaneous vaccination succeeded in suppressing tumor growth in vivo compared to injection. The skin irritation produced by this formulation was tested using an in vitro 3D constructed skin tissue model and an in vivo histological study, which concluded that the formulation did not cause skin irritation. The results suggest that biocompatible IL-mediated dissolution in an oil-based skin penetration enhancer is a promising strategy for transdermal drug delivery.. |
| 26. |
R Sato, K Minamihata, R Wakabayashi, M Goto, N Kamiya, PolyTag: A peptide tag that affords scaffold-less covalent protein assembly catalyzed by microbial transglutaminase., Analytical biochemistry, 10.1016/j.ab.2020.113700, 600, 113700-113700, 2020.07, Assembling proteins in close vicinity to each other provides an opportunity to gain unique function because collaborative and even synergistic functionalities can be expected in an assembled form. There have been a variety of strategies to synthesize functional protein assemblies but site-specific covalent assembly of monomeric protein units without impairing their intrinsic function remains challenging. Herein we report a powerful strategy to design protein assemblies by using microbial transglutaminase (MTG). A serendipitous discovery of self-crosslinking of enhanced green fluorescent protein (EGFP) fused with StrepTag I at the C-terminus revealed that EGFP was assembled through the crosslinking of the Lys (K) residue in the C-terminus of EGFP and the Gln (Q) residue in StrepTag I (AWRHPQFGG). Site-directed mutagenesis of the residues next to the K and Q yielded EGFP assemblies with higher molecular weights. An optimized peptide tag comprised of both K and Q residues (HKRWRHYQRGG) enabled the assembly of different types of proteins of interest (POI) when it was fused to either the N- or C-terminus. The peptide tag that enabled the self-polymerization of the functional POI without a scaffold was designated as a 'PolyTag'.. |
| 27. |
Wahyu Ramadhan, Yuki Ohama, Kosuke Minamihata, Kousuke Moriyama, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya, Redox-responsive functionalized hydrogel marble for the generation of cellular spheroids., Journal of bioscience and bioengineering, 10.1016/j.jbiosc.2020.05.010, 130, 4, 416-423, 2020.07, Liquid marbles (LMs) have recently shown a great promise as microbioreactors to construct self-supported aqueous compartments for chemical and biological reactions. However, the evaporation of the inner aqueous liquid core has limited their application, especially in studying cellular functions. Hydrogels are promising scaffolds that provide a spatial environment suitable for three-dimensional cell culture. Here, we describe the fabrication of redox-responsive hydrogel marbles (HMs) as a three-dimensional cell culture platform. The HMs are prepared by introducing an aqueous mixture of a tetra-thiolated polyethylene glycol (PEG) derivative, thiolated gelatin (Gela-SH), horseradish peroxidase, a small phenolic compound, and human hepatocellular carcinoma cells (HepG2) to the inner aqueous phase of LMs. Eventually, HepG2 cells are encapsulated in the HMs then immersed in culture media, where they proliferate and form cellular spheroids. Experimental results show that the Gela-SH concentration strongly influences the physicochemical and microstructure properties of the HMs. After 6 days in culture, the spheroids were recovered from the HMs by degrading the scaffold, and examination showed that they had reached up to about 180 μm in diameter depending on the Gela-SH concentration, compared with 60 μm in conventional HMs without Gela-SH. After long-term culture (over 12 days), the liver-specific functions (secretion of albumin and urea) and DNA contents of the spheroids cultured in the HMs were elevated compared with those cultured in LMs. These results suggest that the developed HMs can be useful in designing a variety of microbioreactors for tissue engineering applications.. |
| 28. |
Wahyu Ramadhan, Genki Kagawa, Kousuke Moriyama, Rie Wakabayashi, Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Construction of higher-order cellular microstructures by a self-wrapping co-culture strategy using a redox-responsive hydrogel, Scientific reports, 10.1038/s41598-020-63362-4, 10, 1, 2020.05, In this report, a strategy for constructing three-dimensional (3D) cellular architectures comprising viable cells is presented. The strategy uses a redox-responsive hydrogel that degrades under mild reductive conditions, and a confluent monolayer of cells (i.e., cell sheet) cultured on the hydrogel surface peels off and self-folds to wrap other cells. As a proof-of-concept, the self-folding of fibroblast cell sheet was triggered by immersion in aqueous cysteine, and this folding process was controlled by the cysteine concentration. Such folding enabled the wrapping of human hepatocellular carcinoma (HepG2) spheroids, human umbilical vein endothelial cells and collagen beads, and this process improved cell viability, the secretion of metabolites and the proliferation rate of the HepG2 cells when compared with a two-dimensional culture under the same conditions. A key concept of this study is the ability to interact with other neighbouring cells, providing a new, simple and fast method to generate higher-order cellular aggregates wherein different types of cellular components are added. We designated the method of using a cell sheet to wrap another cellular aggregate the ‘cellular Furoshiki’. The simple self-wrapping Furoshiki technique provides an alternative approach to co-culture cells by microplate-based systems, especially for constructing heterogeneous 3D cellular microstructures.. |
| 29. |
Md Rafiqul Islam, Md Raihan Chowdhury, Rie Wakabayashi, Yoshiro Tahara, Noriho Kamiya, Muhammad Moniruzzaman, Masahiro Goto, Choline and amino acid based biocompatible ionic liquid mediated transdermal delivery of the sparingly soluble drug acyclovir, International Journal of Pharmaceutics, 10.1016/j.ijpharm.2020.119335, 582, 2020.05, Transdermal delivery of drugs is more challenging for drugs that are insoluble or sparingly soluble in water and most organic solvents. To overcome this problem, ionic liquid (IL)-mediated ternary systems have been suggested as potential drug carriers. Here, we report potent ternary (IL–EtOH–IPM) systems consisting of biocompatible ILs, ethanol (EtOH), and isopropyl myristate (IPM) that can dissolve a significant amount of the sparingly soluble drug acyclovir (ACV). The ternary systems were optically transparent and thermodynamically stable with a wide range of IL pertinence. An in vitro drug permeation study showed that the ILs in the ternary systems dramatically enhanced ACV permeation into and across the skin. Fourier Transform Infrared spectroscopy of the stratum corneum (sc) after treatment with ternary systems showed that the skin barrier function was reduced by disturbance of the regularly ordered arrangement of corneocytes and modification of the surface properties of the sc during permeation. Histological analysis, and skin irritation studies using a reconstructed human epidermis model showed the safety profile of the ternary system, and there were no significant changes in the structures of the sc, epidermis, and dermis. Therefore, ternary systems containing biocompatible ILs are promising for transdermal delivery of insoluble or sparingly soluble drugs.. |
| 30. |
Md Rafiqul Islam, Md Raihan Chowdhury, Rie Wakabayashi, Noriho Kamiya, Muhammad Moniruzzaman, Masahiro Goto, Ionic liquid-in-oil microemulsions prepared with biocompatible choline carboxylic acids for improving the transdermal delivery of a sparingly soluble drug, Pharmaceutics, 10.3390/pharmaceutics12040392, 12, 4, 2020.04, The transdermal delivery of sparingly soluble drugs is challenging due to of the need for a drug carrier. In the past few decades, ionic liquid (IL)-in-oil microemulsions (IL/O MEs) have been developed as potential carriers. By focusing on biocompatibility, we report on an IL/O ME that is designed to enhance the solubility and transdermal delivery of the sparingly soluble drug, acyclovir. The prepared MEs were composed of a hydrophilic IL (choline formate, choline lactate, or choline propionate) as the non-aqueous polar phase and a surface-active IL (choline oleate) as the surfactant in combination with sorbitan laurate in a continuous oil phase. The selected ILs were all biologically active ions. Optimized pseudo ternary phase diagrams indicated the MEs formed thermodynamically stable, spherically shaped, and nano-sized (90%) with the ME compared with Dulbecco’s phosphate-buffered saline, indicates the biocompatibility of the ME. Therefore, we conclude that IL/O ME may be a promising nano-carrier for the transdermal delivery of sparingly soluble drugs.. |
| 31. |
Masahiko Kobayashi, Jian Xu, Kohei Kakino, Akitsu Masuda, Masato Hino, Naoki Fujimoto, Kosuke Minamihata, Noriho Kamiya, Hiroaki Mon, Hiroshi Iida, Masateru Takahashi, Takahiro Kusakabe, Jae Man Lee, Optimal silkworm larva host for high-level production of Mus musculus IL-4 using a baculovirus expression vector system, Journal of Asia-Pacific Entomology, 10.1016/j.aspen.2019.12.014, 23, 1, 268-273, 2020.04, Interleukine-4 (IL-4) is a cytokine that plays an important role in the immune system and recognized as a biological medicine. Therefore, there is a demand for the production of IL-4 with high performance. The expression of a recombinant IL-4 protein in the prokaryotic system usually results in the formation of an inclusion body. To date, the solution to obtain those active products without the refolding process remains to be established. In this study, we tried to acquire a biologically active recombinant Mus musculus IL-4 (rMmIL-4) using a silkworm-baculovirus expression vector system (silkworm-BEVS). We constructed two recombinant baculoviruses coding rMmIL-4 with the distinct location of affinity purification tags and succeeded in the expression and purification of rMmIL-4 proteins directly without the refolding process. Both purified proteins displayed comparable biological activity to the commercial proteins produced by the E. coli expression system. Besides, we performed screening of silkworm strains to seek optimal hosts for the mass-production of rMmIL-4. Intriguingly, we found that some silkworm strains showed significantly higher secretion levels of rMmIL-4 in silkworm sera. Our study provides meaningful insights into the industrial-scale production of rMmIL-4 with high productivity for pharmaceutical applications in the future.. |
| 32. |
Mari Takahara, Noriho Kamiya, Synthetic Strategies for Artificial Lipidation of Functional Proteins, Chemistry - A European Journal, 10.1002/chem.201904568, 26, 21, 4645-4655, 2020.04, Biosynthesis of natural lipidated proteins is linked to important signal pathways, and therefore analyzing protein lipidation is crucial for understanding cellular functions. Artificial lipidation of proteins has attracted attention in recent decades as it allows modulation of the amphiphilic nature of the protein of interest, and is used in the design of drug-delivery systems containing antibodies anchored on a lipid bilayer carrier. However, the intrinsic hydrophobicity of lipids makes the synthesis of lipid–protein conjugates challenging with respect to the yield and selectivity of the lipidation. In this Minireview, the development of chemical and enzymatic synthetic strategies for the preparation of a range of lipid–protein conjugates that do not compromise the functions of the proteins are discussed as well as applications of the conjugates.. |
| 33. |
Wahyu Ramadhan, Genki Kagawa, Kousuke Moriyama, Rie Wakabayashi, Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Construction of higher-order cellular microstructures by a self-wrapping co-culture strategy using a redox-responsive hydrogel., Scientific reports, 10.1038/s41598-020-63362-4, 10, 1, 6710-6710, 2020.04, In this report, a strategy for constructing three-dimensional (3D) cellular architectures comprising viable cells is presented. The strategy uses a redox-responsive hydrogel that degrades under mild reductive conditions, and a confluent monolayer of cells (i.e., cell sheet) cultured on the hydrogel surface peels off and self-folds to wrap other cells. As a proof-of-concept, the self-folding of fibroblast cell sheet was triggered by immersion in aqueous cysteine, and this folding process was controlled by the cysteine concentration. Such folding enabled the wrapping of human hepatocellular carcinoma (HepG2) spheroids, human umbilical vein endothelial cells and collagen beads, and this process improved cell viability, the secretion of metabolites and the proliferation rate of the HepG2 cells when compared with a two-dimensional culture under the same conditions. A key concept of this study is the ability to interact with other neighbouring cells, providing a new, simple and fast method to generate higher-order cellular aggregates wherein different types of cellular components are added. We designated the method of using a cell sheet to wrap another cellular aggregate the 'cellular Furoshiki'. The simple self-wrapping Furoshiki technique provides an alternative approach to co-culture cells by microplate-based systems, especially for constructing heterogeneous 3D cellular microstructures.. |
| 34. |
Md Rafiqul Islam, Md Raihan Chowdhury, Rie Wakabayashi, Noriho Kamiya, Muhammad Moniruzzaman, Masahiro Goto, Ionic Liquid-In-Oil Microemulsions Prepared with Biocompatible Choline Carboxylic Acids for Improving the Transdermal Delivery of a Sparingly Soluble Drug., Pharmaceutics, 10.3390/pharmaceutics12040392, 12, 4, 2020.04, The transdermal delivery of sparingly soluble drugs is challenging due to of the need for a drug carrier. In the past few decades, ionic liquid (IL)-in-oil microemulsions (IL/O MEs) have been developed as potential carriers. By focusing on biocompatibility, we report on an IL/O ME that is designed to enhance the solubility and transdermal delivery of the sparingly soluble drug, acyclovir. The prepared MEs were composed of a hydrophilic IL (choline formate, choline lactate, or choline propionate) as the non-aqueous polar phase and a surface-active IL (choline oleate) as the surfactant in combination with sorbitan laurate in a continuous oil phase. The selected ILs were all biologically active ions. Optimized pseudo ternary phase diagrams indicated the MEs formed thermodynamically stable, spherically shaped, and nano-sized (90%) with the ME compared with Dulbecco's phosphate-buffered saline, indicates the biocompatibility of the ME. Therefore, we conclude that IL/O ME may be a promising nano-carrier for the transdermal delivery of sparingly soluble drugs.. |
| 35. |
Yuya Hirakawa, Hiroshi Ueda, Rie Wakabayashi, Noriho Kamiya, Masahiro Goto, A novel binary supercooled liquid formulation for transdermal drug delivery, Biological and Pharmaceutical Bulletin, 10.1248/bpb.b19-00642, 43, 3, 393-398, 2020.03, The aim of this study was to prepare binary supercooled liquid (SCL) by intermolecular interaction and apply this formulation to transdermal drug delivery. Ketoprofen (KET) and ethenzamide (ETH) were selected as binary SCL component. Thermal analysis of physical mixtures of KET and ETH showed decreases in melting points and glass transition below room temperature, thereby indicating formation of KET–ETH SCL. Intermolecular interactions between KET and ETH in the SCL were evaluated from Fourier transform (FT)-IR spectra. KET–ETH SCL maintained SCL state at 25°C with silica gel over 31d and at 40°C/89% relative humidity (RH) over 7d. KET SCL and KET–ETH SCL showed similar permeability of KET for hairless mice skin, which was two-fold higher than that of KET aqueous suspension. Our findings suggest that the SCL state could enhance the skin permeation of drugs and the binary SCL formed by intermolecular interaction could also improve the stability of the SCL. The binary SCL system could become a new drug form for transdermal drug delivery.. |
| 36. |
Momoko Kitaoka, Wei Xiao, Qingliang Kong, Yoshiro Tahara, Noriho Kamiya, Masahiro Goto, A solid-in-oil nanodispersion system for transcutaneous immunotherapy of cow’s milk allergies, Pharmaceutics, 10.3390/pharmaceutics12030205, 12, 3, 2020.03, An allergy to cow’s milk proteins is the most common food allergy in infants and toddlers. Conventional oral immunotherapy for cow’s milk allergies requires hospital admission due to the risk of severe allergic reactions, including anaphylaxis. Therefore, a simpler and safer immunotherapeutic method is desirable. We examined transcutaneous immunotherapy with a solid-in-oil (S/O) system. In the S/O system, nano-sized particles of proteins are dispersed in an oil-vehicle with the assistance of nonionic surfactants. In the present study, the S/O system enhanced the skin permeation of the allergen molecule β-lactoglobulin (BLG), as compared with a control PBS solution. The patches containing BLG in the S/O nanodispersion skewed the immune response in the allergy model mice toward T helper type 1 immunity, indicating the amelioration of allergic symptoms. This effect was more pronounced when the immunomodulator resiquimod (R-848) was included in the S/O system.. |
| 37. |
Dani Permana, Kosuke Minamihata, Ryo Sato, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya, Linear Polymerization of Protein by Sterically Controlled Enzymatic Cross-Linking with a Tyrosine-Containing Peptide Loop, ACS Omega, 10.1021/acsomega.9b04163, 5, 10, 5160-5169, 2020.03, The structure of a protein complex needs to be controlled appropriately to maximize its functions. Herein, we report the linear polymerization of bacterial alkaline phosphatase (BAP) through the site-specific cross-linking reaction catalyzed by Trametes sp. laccase (TL). We introduced a peptide loop containing a tyrosine (Y-Loop) to BAP, and the Y-Looped BAP was treated with TL. The Y-Looped BAP formed linear polymers, whereas BAP fused with a C-terminal peptide containing a tyrosine (Y-tag) showed an irregular shape after TL treatment. The sterically confined structure of the Y-Loop could be responsible for the formation of linear BAP polymers. TL-catalyzed copolymerization of Y-Looped BAP and a Y-tagged chimeric antibody-binding protein, pG2pA-Y, resulted in the formation of linear bifunctional protein copolymers that could be employed as protein probes in an enzyme-linked immunosorbent assay (ELISA). Copolymers comprising Y-Looped BAP and pG2pA-Y at a molar ratio of 100:1 exhibited the highest signal in the ELISA with 26- and 20-fold higher than a genetically fused chimeric protein, BAP-pG2pA-Y, and its polymeric form, respectively. This result revealed that the morphology of the copolymers was the most critical feature to improve the functionality of the protein polymers as detection probes, not only for immunoassays but also for other diagnostic applications.. |
| 38. |
Qingliang Kong, Momoko Kitaoka, Rie Wakabayashi, Yoshiro Tahara, Noriho Kamiya, Masahiro Goto, Solid-in-oil nanodispersions for transcutaneous immunotherapy of Japanese cedar pollinosis, Pharmaceutics, 10.3390/pharmaceutics12030240, 12, 3, 2020.03, Japanese cedar pollinosis (JCP) is a common affliction caused by an allergic reaction to cedar pollen and is considered a disease of national importance in Japan. Antigen-specific immunotherapy (AIT) is the only available curative treatment for JCP. However, low compliance and persistence have been reported among patients subcutaneously or sublingually administered AIT comprising a conventional antigen derived from a pollen extract. To address these issues, many research studies have focused on developing a safer, simpler, and more effective AIT for JCP. Here, we review the novel antigens that have been developed for JCP AIT, discuss their different administration routes, and present the effects of anti-allergy treatment. Then, we describe a new form of AIT called transcutaneous immunotherapy (TCIT) and its solid-in-oil (S/O) nanodispersion formulation, which is a promising antigen delivery system. Finally, we discuss the applications of S/O nanodispersions for JCP TCIT. In this context, we predict that TCIT delivery by using a S/O nanodispersion loaded with novel antigens may offer an easier, safer, and more effective treatment option for JCP patients.. |
| 39. |
Dani Permana, Kosuke Minamihata, Ryo Sato, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya, Linear Polymerization of Protein by Sterically Controlled Enzymatic Cross-Linking with a Tyrosine-Containing Peptide Loop., ACS omega, 10.1021/acsomega.9b04163, 5, 10, 5160-5169, 2020.03, The structure of a protein complex needs to be controlled appropriately to maximize its functions. Herein, we report the linear polymerization of bacterial alkaline phosphatase (BAP) through the site-specific cross-linking reaction catalyzed by Trametes sp. laccase (TL). We introduced a peptide loop containing a tyrosine (Y-Loop) to BAP, and the Y-Looped BAP was treated with TL. The Y-Looped BAP formed linear polymers, whereas BAP fused with a C-terminal peptide containing a tyrosine (Y-tag) showed an irregular shape after TL treatment. The sterically confined structure of the Y-Loop could be responsible for the formation of linear BAP polymers. TL-catalyzed copolymerization of Y-Looped BAP and a Y-tagged chimeric antibody-binding protein, pG2pA-Y, resulted in the formation of linear bifunctional protein copolymers that could be employed as protein probes in an enzyme-linked immunosorbent assay (ELISA). Copolymers comprising Y-Looped BAP and pG2pA-Y at a molar ratio of 100:1 exhibited the highest signal in the ELISA with 26- and 20-fold higher than a genetically fused chimeric protein, BAP-pG2pA-Y, and its polymeric form, respectively. This result revealed that the morphology of the copolymers was the most critical feature to improve the functionality of the protein polymers as detection probes, not only for immunoassays but also for other diagnostic applications.. |
| 40. |
Qingliang Kong, Momoko Kitaoka, Rie Wakabayashi, Yoshiro Tahara, Noriho Kamiya, Masahiro Goto, Solid-in-Oil Nanodispersions for Transcutaneous Immunotherapy of Japanese Cedar Pollinosis., Pharmaceutics, 10.3390/pharmaceutics12030240, 12, 3, 2020.03, Japanese cedar pollinosis (JCP) is a common affliction caused by an allergic reaction to cedar pollen and is considered a disease of national importance in Japan. Antigen-specific immunotherapy (AIT) is the only available curative treatment for JCP. However, low compliance and persistence have been reported among patients subcutaneously or sublingually administered AIT comprising a conventional antigen derived from a pollen extract. To address these issues, many research studies have focused on developing a safer, simpler, and more effective AIT for JCP. Here, we review the novel antigens that have been developed for JCP AIT, discuss their different administration routes, and present the effects of anti-allergy treatment. Then, we describe a new form of AIT called transcutaneous immunotherapy (TCIT) and its solid-in-oil (S/O) nanodispersion formulation, which is a promising antigen delivery system. Finally, we discuss the applications of S/O nanodispersions for JCP TCIT. In this context, we predict that TCIT delivery by using a S/O nanodispersion loaded with novel antigens may offer an easier, safer, and more effective treatment option for JCP patients.. |
| 41. |
Rahman Md Moshikur, Md Raihan Chowdhury, Rie Wakabayashi, Yoshiro Tahara, Noriho Kamiya, Muhammad Moniruzzaman, Masahiro Goto, Ionic liquids with N-methyl-2-pyrrolidonium cation as an enhancer for topical drug delivery
Synthesis, characterization, and skin-penetration evaluation, Journal of Molecular Liquids, 10.1016/j.molliq.2019.112166, 299, 2020.02, The development of non-toxic ionic liquid-based active pharmaceutical ingredients (IL-APIs) for effective topical drug delivery is still challenging. The properties of IL-APIs can be boosted up by selecting potential biocompatible cations. Here, we introduced N-methyl-2-pyrrolidone (NMP) as a potent biocompatible counter ion to prepare ionic liquefied drugs for topical drug delivery. The cytotoxicity of NMP cation was investigated using mammalian cell lines (HepG2, NIH3T3 and L929 cells) and compared with conventional IL-forming cations. The synthesized NMP cation has lower toxicity than that of conventional IL-forming cations. The NMP cation showed at least 3.6, 15.2 and 58.9 times lower toxicity than that of conventional imidazolium, ammonium and phosphonium cations, respectively. The synthesized NMP-based ionic liquid (NMP-IL) was characterized using 1H & 13C NMR, FT-IR, DSC and TGA. NMP-IL showed better physico-thermal stability, enhanced skin penetration, and enriched drug accumulation 2.6 times higher than that of IL [Cho][Ibu] in the target tissue. These results suggested that NMP cation based API-IL can be an effective biocompatible formulation for topical drug delivery by accumulating active drugs in the skin.. |
| 42. |
Shuto Kozaka, Yoshiro Tahara, Rie Wakabayashi, Takahiro Nakata, Taro Ueda, Noriho Kamiya, Masahiro Goto, Transcutaneous Cancer Vaccine Using a Reverse Micellar Antigen Carrier, Molecular pharmaceutics, 10.1021/acs.molpharmaceut.9b01104, 17, 2, 645-655, 2020.02, Skin dendritic cells (DCs) such as Langerhans cells and dermal dendritic cells have a pivotal role in inducing antigen-specific immunity; therefore, transcutaneous cancer vaccines are a promising strategy to prophylactically prevent the onset of a variety of diseases, including cancers. The largest obstacle to delivering antigen to these skin DC subsets is the barrier function of the stratum corneum. Although reverse micellar carriers are commonly used to enhance skin permeability to hydrophilic drugs, the transcutaneous delivery of antigen, proteins, or peptides has not been achieved to date because of the large molecular weight of drugs. To achieve effective antigen delivery to skin DCs, we developed a novel strategy using a surfactant as a skin permeation enhancer in a reverse micellar carrier. In this study, glyceryl monooleate (MO) was chosen as a skin permeation enhancer, and the MO-based reverse micellar carrier enabled the successful delivery of antigen to Langerhans cells and dermal dendritic cells. Moreover, transcutaneous vaccination with the MO-based reverse micellar carrier significantly inhibited tumor growth, indicating that it is a promising vaccine platform against tumors.. |
| 43. |
Shuto Kozaka, Yoshiro Tahara, Rie Wakabayashi, Takahiro Nakata, Taro Ueda, Noriho Kamiya, Masahiro Goto, Transcutaneous Cancer Vaccine Using a Reverse Micellar Antigen Carrier., Molecular pharmaceutics, 10.1021/acs.molpharmaceut.9b01104, 17, 2, 645-655, 2020.02, Skin dendritic cells (DCs) such as Langerhans cells and dermal dendritic cells have a pivotal role in inducing antigen-specific immunity; therefore, transcutaneous cancer vaccines are a promising strategy to prophylactically prevent the onset of a variety of diseases, including cancers. The largest obstacle to delivering antigen to these skin DC subsets is the barrier function of the stratum corneum. Although reverse micellar carriers are commonly used to enhance skin permeability to hydrophilic drugs, the transcutaneous delivery of antigen, proteins, or peptides has not been achieved to date because of the large molecular weight of drugs. To achieve effective antigen delivery to skin DCs, we developed a novel strategy using a surfactant as a skin permeation enhancer in a reverse micellar carrier. In this study, glyceryl monooleate (MO) was chosen as a skin permeation enhancer, and the MO-based reverse micellar carrier enabled the successful delivery of antigen to Langerhans cells and dermal dendritic cells. Moreover, transcutaneous vaccination with the MO-based reverse micellar carrier significantly inhibited tumor growth, indicating that it is a promising vaccine platform against tumors.. |
| 44. |
Rie Wakabayashi, Wahyu Ramadhan, Kousuke Moriyama, Masahiro Goto, Noriho Kamiya, Poly(ethylene glycol)-based biofunctional hydrogels mediated by peroxidase-catalyzed cross-linking reactions, Polymer Journal, 10.1038/s41428-020-0344-7, 2020.01, Biofunctional hydrogels prepared by a peroxidase, especially horseradish peroxidase (HRP), serve as an excellent class of materials or platform for the development of cellular scaffolds because their biocompatibility and mild and tunable reaction conditions provide them with desirable properties. In this focus review, we summarize our decade of research into HRP-mediated fabrication of biofunctional hydrogels and their applications, in particular cell culture scaffolds. A brief overview of potential substrates employed in HRP and improvement of the HRP hydrogelation system from the initial step until the hydrogen peroxide removal stage in an effort to meet environmental standards is discussed. We highlight our system and describe its biocompatibility and ability to functionalize molecules to support biofabrication by increasing cellular adhesiveness, retaining growth factor affinity, and finally accelerating the formation of two- and three-dimensional multicellular architectures. In the last section, we outline the adoption of hydrogelation as a self-standing, compartmentalized reaction system, i.e., the use of hydrogel marble to conduct cell-free biosynthesis. We believe that this HRP-mediated hydrogel system offers great potential not only as a cell culture scaffold but also for various biomedical applications.. |
| 45. |
R. Sato, K. Minamihata, R. Wakabayashi, M. Goto, N. Kamiya, PolyTag
A peptide tag that affords scaffold-less covalent protein assembly catalyzed by microbial transglutaminase, Analytical Biochemistry, 10.1016/j.ab.2020.113700, 2020.01, Assembling proteins in close vicinity to each other provides an opportunity to gain unique function because collaborative and even synergistic functionalities can be expected in an assembled form. There have been a variety of strategies to synthesize functional protein assemblies but site-specific covalent assembly of monomeric protein units without impairing their intrinsic function remains challenging. Herein we report a powerful strategy to design protein assemblies by using microbial transglutaminase (MTG). A serendipitous discovery of self-crosslinking of enhanced green fluorescent protein (EGFP) fused with StrepTag I at the C-terminus revealed that EGFP was assembled through the crosslinking of the Lys (K) residue in the C-terminus of EGFP and the Gln (Q) residue in StrepTag I (AWRHPQFGG). Site-directed mutagenesis of the residues next to the K and Q yielded EGFP assemblies with higher molecular weights. An optimized peptide tag comprised of both K and Q residues (HKRWRHYQRGG) enabled the assembly of different types of proteins of interest (POI) when it was fused to either the N- or C-terminus. The peptide tag that enabled the self-polymerization of the functional POI without a scaffold was designated as a ‘PolyTag’.. |
| 46. |
Qingliang Kong, Momoko Kitaoka, Yoshiro Tahara, Rie Wakabayashi, Noriho Kamiya, Masahiro Goto, Solid-in-oil nanodispersions for intranasal vaccination
Enhancement of mucosal and systemic immune responses, International Journal of Pharmaceutics, 10.1016/j.ijpharm.2019.118777, 572, 2019.12, En masse vaccination is a promising strategy for combatting infectious diseases. Intranasal vaccination is a viable route of mass vaccination, and it could be performed easily via needle-free administration. However, it is not widely used because it tends not to evoke sufficient immunity. The aim of the present study was to improve the performance of intranasal vaccination by extending the amount of time that administered antigens remain in the nasal cavity, and enhancing immune responses via a nanocarrier-based adjuvant. A simple and safe solid-in-oil (S/O) system was investigated as a nanocarrier in intranasal vaccination. S/O nanodispersions are oil-based dispersions of antigens coated with surfactants. Because of the mucoadhesive capacities of surfactant and oil they have high potential to extend the amount of time that administered antigens remain in the nasal cavity, and can induce strong immune responses due to a nanocarrier-based adjuvant effect. In nasal absorption experiments antigens administered intranasally via S/O nanodispersions remained in the nasal cavity longer and induced strong mucosal and systemic immune responses. Histopathology analysis indicated that S/O nanodispersions did not modify the nasal epithelium or cilia, suggesting non-toxicity of the carrier. These results indicate the potential of intranasal vaccination using S/O nanodispersions for future vaccination.. |
| 47. |
A novel binary supercooled liquid formulation for transdermal drug delivery.. |
| 48. |
Solid-in-oil nanodispersions for intranasal vaccination: Enhancement of mucosal and systemic immune responses.. |
| 49. |
Mari Takahara, Noriho Kamiya, Synthetic Strategies for Artificial Lipidation of Functional Proteins, Chemistry-A European Journal, 10.1002/chem.201904568, Accepted, 2019.12. |
| 50. |
Qingliang Kong, Kouki Higasijima, Rie Wakabayashi, Yoshiro Tahara, Momoko Kitaoka, Hiroki Obayashi, Yanting Hou, Noriho Kamiya, Masahiro Goto, Transcutaneous delivery of immunomodulating pollen extract-galactomannan conjugate by solid-in-oil nanodispersions for pollinosis immunotherapy, Pharmaceutics, 10.3390/pharmaceutics11110563, 11, 11, 2019.11, Japanese cedar pollinosis is a type I allergic disease and has already become a major public health problem in Japan. Conventional subcutaneous immunotherapy (SCIT) and sublingual immunotherapy (SLIT) cannot meet patients’ needs owing to the side effects caused by both the use of conventional whole antigen molecules in the pollen extract and the administration routes. To address these issues, a surface-modified antigen and transcutaneous administration route are introduced in this research. First, the pollen extract (PE) was conjugated to galactomannan (PE-GM) to mask immunoglobulin E (IgE)-binding epitopes in the PE to avoid side effects. Second, as a safer alternative to SCIT and SLIT, transcutaneous immunotherapy (TCIT) with a solid-in-oil (S/O) nanodispersion system carrying PE-GM was proposed. Hydrophilic PE-GM was efficiently delivered through mouse skin using S/O nanodispersions, reducing the antibody secretion and modifying the type 1 T helper (Th1)/ type 2 T helper (Th2) balance in the mouse model, thereby demonstrating the potential to alleviate Japanese cedar pollinosis.. |
| 51. |
Yuya Hirakawa, Hiroshi Ueda, Tetsuya Miyano, Noriho Kamiya, Masahiro Goto, New insight into transdermal drug delivery with supersaturated formulation based on co-amorphous system, International Journal of Pharmaceutics, 10.1016/j.ijpharm.2019.118582, 569, 2019.10, The objective of this study was to prepare a supersaturated formulation based on formation of a co-amorphous system of a drug and a coformer in order to enhance skin permeation. Atenolol (ATE) and urea (URE) were used as the model drug and the coformer, respectively. Thermal analysis of physical mixtures of ATE and URE showed decreases in the melting points and the formation of a co-amorphous system which was in a supercooled liquid state because of a low glass transition temperature. Supersaturated solutions of ATE and URE at different molar ratios in polyethylene glycol 400 (PEG400) were prepared. The precipitations were observed under storage at 25 °C for all formulations except for ATE-URE at 1:8 molar ratio which remained in the supersaturated state for 2 months. 1H NMR analysis confirmed the interactions between ATE and URE in PEG400. The ATE-URE supersaturated formulation showed higher permeability for mice skin than that of ATE saturated formulation, which was superior to the expected permeability from the degree of supersaturation. We concluded that co-amorphous based supersaturated formulation offers much promise for transdermal drug delivery.. |
| 52. |
Transcutaneous Delivery of Immunomodulating Pollen Extract-Galactomannan Conjugate by Solid-in-Oil Nanodispersions for Pollinosis Immunotherapy.. |
| 53. |
Md Korban Ali, Rahman Md Moshikur, Rie Wakabayashi, Yoshiro Tahara, Muhammad Moniruzzaman, Noriho Kamiya, Masahiro Goto, Synthesis and characterization of choline–fatty-acid-based ionic liquids
A new biocompatible surfactant, Journal of Colloid And Interface Science, 10.1016/j.jcis.2019.04.095, 551, 72-80, 2019.09, Ionic liquid (IL)surfactants have attracted great interest as promising substitutes for conventional surfactants owing to their exceptional and favorable physico-chemical properties. However, most IL surfactants are not eco-friendly and form unstable micelles, even when using a high concentration of the surfactant. In this study, we prepared a series of halogen-free and biocompatible choline–fatty-acid-based ILs with different chain lengths and degrees of saturation, and we then investigated their micellar properties in aqueous solutions. Characterization of the synthesized surface-active ILs (SAILs)was performed by 1 H and 13 C nuclear magnetic resonance spectroscopy, Fourier transform infrared spectroscopy, differential scanning calorimetry, and elemental analysis. The surface-active properties of the SAILs were investigated by tensiometry, conductometry, and dynamic light scattering measurements. The critical micelle concentration of the SAILs was found to be 2–4 times lower than those of conventional surfactants. The thermodynamic properties of micellization (ΔG 0 m , ΔH 0 m , and ΔS 0 m )indicate that the micellization process of the SAILs is spontaneous, stable, and entropy-driven at room temperature. The cytotoxicity of the SAILs was evaluated using mammalian cell line NIH 3T3. Importantly, [Cho][Ole]shows lower toxicity than the analogous ILs with conventional surfactants. These results clearly suggest that these environmentally friendly SAILs can be used as a potential alternative to conventional ILs for various purposes, including biological applications.. |
| 54. |
Safrina Dyah Hardiningtyas, Seiya Nagao, Emiko Yamamoto, Nana Shirakigawa, Rie Wakabayashi, Masahiro Goto, Hiroyuki Ijima, Noriho Kamiya, A nano-sized gel-in-oil suspension for transcutaneous protein delivery, International Journal of Pharmaceutics, 10.1016/j.ijpharm.2019.118495, 567, 2019.08, We developed a new oil-based delivery system for transdermal protein delivery, a gel-in-oil (G/O) nanosuspension, where gelatin-based hydrogel was coated with hydrophobic surfactants. The high entrapment efficiency of a model protein, phycocyanin (PC), into nano-sized gelatin hydrogel particles was achieved. Spectroscopic evaluation of PC suggested that the G/O nanosuspension could retain the functional form of PC in isopropyl myristate. In vitro skin permeation studies showed that the G/O nanosuspension facilitated the delivery of PC through the stratum corneum of Yucatan micropig skin.. |
| 55. |
Safrina Dyah Hardiningtyas, Seiya Nagao, Emiko Yamamoto, Nana Shirakigawa, Rie Wakabayashi, Masahiro Goto, Hiroyuki Ijima, Noriho Kamiya, A nano-sized gel-in-oil suspension for transcutaneous protein delivery, 10.1016/j.ijpharm.2019.118495, 567, 118495-118495, 2019.08. |
| 56. |
Yurie Kinoshita, Jian Xu, Akitsu Masuda, Kosuke Minamihata, Noriho Kamiya, Hiroaki Mon, Ryosuke Fujita, Takahiro Kusakabe, Man Lee, Expression and purification of biologically active human granulocyte-macrophage colony stimulating factor (hGM-CSF) using silkworm-baculovirus expression vector system, Protein Expression and Purification, 10.1016/j.pep.2019.03.010, 159, 69-74, 2019.07, Human granulocyte-macrophage colony stimulating factor (hGM-CSF) is a hematopoietic growth factor. It is widely employed as a therapeutic agent targeting neutropenia in cancer patients undergoing chemotherapy and in patients with AIDS or after bone marrow transplantation. In this study, we constructed the recombinant baculoviruses for the expression of recombinant hGM-CSF (rhGM-CSF) with two small affinity tags (His-tag and Strep-tag) at the N or C-terminus. Compared to N-tagged rhGM-CSF, C-tagged rhGM-CSF was highly recovered from silkworm hemolymph. The purified rhGM-CSF proteins migrated as a diffuse band and were confirmed to hold N-glycosylations. A comparable activity was achieved when commercial hGM-CSF was tested as a control. Considering the high price of hGM-CSF in the market, our results and strategies using silkworm-baculovirus system can become a great reference for mass production of the active rhGM-CSF at a lower cost.. |
| 57. |
Expression and purification of biologically active human granulocyte-macrophage colony stimulating factor (hGM-CSF) using silkworm-baculovirus expression vector system.. |
| 58. |
Wahyu Ramadhan, Genki Kagawa, Yusei Hamada, Kousuke Moriyama, Rie Wakabayashi, Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Enzymatically Prepared Dual Functionalized Hydrogels with Gelatin and Heparin to Facilitate Cellular Attachment and Proliferation, ACS Applied Bio Materials, 10.1021/acsabm.9b00275, 2, 6, 2600-2609, 2019.06, Biologically active artificial scaffolds for cell seeding are developed by mimicking extracellular matrices using synthetic materials. Here, we propose a feasible approach employing biocatalysis to integrate natural components, that is, gelatin and heparin, into a synthetic scaffold, namely a polyethylene glycol (PEG)-based hydrogel. Initiation of horseradish peroxidase-mediated redox reaction enabled both hydrogel formation of tetra-thiolated PEG via disulfide linkage and incorporation of chemically thiolated gelatin (Gela-SH) and heparin (Hepa-SH) into the polymeric network. We found that the compatibility of the type of gelatin with heparin was crucial for the hydrogelation process. Alkaline-treated gelatin exhibited superior performance over acid-treated gelatin to generate dual functionality in the resultant hydrogel originating from the two natural biopolymers. The Gela-SH/Hepa-SH dual functionalized PEG-based hydrogel supported both cellular attachment and binding of basic fibroblast growth factor (bFGF) under cell culture conditions, which increased the proliferation and phenotype transformation of NIH3T3 cells cultured on the hydrogel. Inclusion of bFGF and a commercial growth factor cocktail in hydrogel matrices effectively enhanced cell spreading and confluency of both NIH3T3 cells and HUVECs, respectively, suggesting a potential method to design artificial scaffolds containing active growth factors.. |
| 59. |
Md Raihan Chowdhury, Rahman Md Moshikur, Rie Wakabayashi, Yoshiro Tahara, Noriho Kamiya, Muhammad Moniruzzaman, Masahiro Goto, In vivo biocompatibility, pharmacokinetics, antitumor efficacy, and hypersensitivity evaluation of ionic liquid-mediated paclitaxel formulations, International Journal of Pharmaceutics, 10.1016/j.ijpharm.2019.05.020, 565, 219-226, 2019.06, In order to prevent common hypersensitivity reactions to paclitaxel injections (Taxol), we previously reported an ionic liquid-mediated paclitaxel (IL-PTX)formulation with small particle size and narrow size distribution. The preliminary work showed high PTX solubility in the IL, and the formulation demonstrated similar antitumor activity to Taxol, while inducing a smaller hypersensitivity effect in in vitro cell experiments. In this study, the stability of the IL-PTX formulation was monitored by quantitative HPLC analysis, which showed that IL-PTX was more stable at 4 °C than at room temperature. The in vivo study showed that the IL-PTX formulation could be used in a therapeutic application as a biocompatible component of a drug delivery system. To assess the in-vivo biocompatibility, IL or IL-mediated formulations were administered intravenously by maintaining physiological buffered conditions (neutral pH and isotonic salt concentration). From in vivo pharmacokinetics data, the IL-PTX formulation was found to have a similar systemic circulation time and slower elimination rate compared to cremophor EL mediated paclitaxel (CrEL-PTX). Furthermore, in vivo antitumor and hypersensitivity experiments in C57BL/6 mice revealed that IL-PTX had similar antitumor activity to CrEL-PTX, but a significantly smaller hypersensitivity effect compared with CrEL-PTX. Therefore, the IL-mediated formulation has potential to be an effective and safe drug delivery system for PTX.. |
| 60. |
Complementary interaction with peptide amphiphiles guides size-controlled assembly of small molecules for intracellular delivery. |
| 61. |
Development of a novel ionic liquid-curcumin complex to enhance its solubility, stability, and activity. |
| 62. |
Permana D, Minamihata K, Tatsuke T, Lee JM, Kusakabe T, Goto M, Kamiya N, Polymerization of Horseradish Peroxidase by a Laccase-Catalyzed Tyrosine Coupling Reaction., Biotechnology journal, 10.1002/biot.201800531, 14, 6, e1800531, 2019.06. |
| 63. |
Mari Takahara, Rie Wakabayashi, Naoki Fujimoto, Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Enzymatic Cell-Surface Decoration with Proteins using Amphiphilic Lipid-Fused Peptide Substrates, Chemistry - A European Journal, 10.1002/chem.201900370, 25, 30, 7315-7321, 2019.05, Lipid modification of proteins plays a significant role in the activation of cellular signals such as proliferation. Thus, the demand for lipidated proteins is rising. However, getting a high yield and purity of lipidated proteins has been challenging. We developed a strategy for modifying proteins with a wide variety of synthetic lipids using microbial transglutaminase (MTG), which catalyzes the cross-linking reaction between a specific glutamine (Q) in a protein and lysine (K) in the lipid-fused peptide. The synthesized lipid substrates (lipid: fatty acids, tocopherol, lithocholic acid, cholesterol) was successfully conjugated to a protein fused with LLQG (Q-tagged protein) by an MTG reaction, yielding 90 % conversion of the Q-tagged protein in a lipidated form. The purified lipid–protein conjugates were used for labeling the cell membrane in vitro, resulting in best-anchoring ability of cholesterol modification. Furthermore, in situ cell-surface decoration with the protein was established in a simple manner: subjection of cells to a mixture of cholesterol-fused peptides, Q-tagged proteins and MTG.. |
| 64. |
Patmawati, Kosuke Minamihata, Tsuneyuki Tatsuke, Man Lee, Takahiro Kusakabe, Noriho Kamiya, Functional horseradish peroxidase−streptavidin chimeric proteins prepared using a silkworm-baculovirus expression system for diagnostic purposes, Journal of Biotechnology, 10.1016/j.jbiotec.2019.03.007, 297, 28-31, 2019.05, Rapid, convenient, sensitive detection methods are of the utmost importance in analytical tools. Enzyme-based signal amplification using horseradish peroxidase (HRP) is commonly implemented in clinical diagnostics kits based on enzyme-linked immunosorbent assay (ELISA), by which the limit of detection is greatly improved. Herein we report the design and preparation of recombinant fusion proteins comprising HRP and streptavidin (Stav), in which HRP was fused to either the N- or C-terminus of Stav ((HRP)
4
–Stav or Stav–(HRP)
4
, respectively) using a baculovirus-silkworm expression system. Both (HRP)
4
–Stav and Stav–(HRP)
4
were secreted in the apo form but they were easily converted to the holo form and activated by simple incubation with hemin overnight at 4 °C. The activated (HRP)
4
–Stav and Stav–(HRP)
4
could be combined with a commercial biotinylated anti-OVA IgG antibody to detect ovalbumin (OVA) as the antigen in ELISA. The enzymatic activity of (HRP)
4
–Stav was twofold higher than that of Stav–(HRP)
4
, and the sensitivity of (HRP)
4
-Stav in ELISA was higher than that of a commercial HRP–Stav chemical conjugate. The successful use of (HRP)
4
–Stav chimeric protein as a molecular probe in ELISA shows that the baculovirus-silkworm expression system is promising to produce enzyme–Stav conjugates to substitute for those prepared by chemical methods.. |
| 65. |
Yovita Djohan, Tomoki Azukizawa, Patmawati, Kotaro Sakai, Yuki Yano, Fumiya Sato, Ryoji Takahashi, Masafumi Yohda, Mizuo Maeda, Noriho Kamiya, Tamotsu Zako, Molecular chaperone prefoldin-assisted biosynthesis of gold nanoparticles with improved size distribution and dispersion, Biomaterials Science, 10.1039/c8bm01026a, 7, 5, 1801-1804, 2019.05, Here we report a novel aspect of molecular chaperone prefoldin (PFD) as a biomaterial in the biocatalytic synthesis of gold nanoparticles (AuNPs) using glycerol dehydrogenase (GLD). We found that PFD could inhibit the aggregation of AuNPs during the biosynthesis, leading to the formation of AuNPs with controlled size distribution.. |
| 66. |
Uju, E. Prasetyaningsih, J. Santoso, N. Kamiya, T. Oshima, Preparation and characterization of semi-refined carrageenan from Kappaphycus alvarezii seaweed bleached by Peracetic Acid, 3rd EMBRIO International Workshop on Marine Biodiversity: Understanding, Utilization, Conservation, EIW 2018
IOP Conference Series: Earth and Environmental Science, 10.1088/1755-1315/278/1/012077, 278, 1, 2019.05, Semi-refined carrageenan (SRC) is one of the products from Kappaphycus alvarezii, which has the potential to be developed in Indonesia. However, unbleached seaweed will produce SRC with light brown colour due to natural seaweed pigments. Peracetic acid (PAA) is a strong oxidizing agent that has the potential to be used as a bleaching agent in the SRC production. This research aimed to study the effects of the PAA as a bleaching agent on the characteristics of SRC from K. alvarezii. Chopped seaweed was heated in 10% KOH at 80°C for 30 minutes. Bleaching was carried out at room temperature for 90 minutes using PAA at a concentration of 0.5%, 1.5%, and 2.5% (w/w) or using 1.5% (w/w) of sodium hypochlorite. Bleaching using PAA produced SRC whiter than without bleaching and bleaching using sodium hypochlorite. SRC brightness increased with increasing concentrations of PAA and while the yield reduced, as well as the viscosity, gel strength, sulfate content and ash content. The best concentration of PAA was 0.5%. At best, PAA concentration bleaching produced white SRC flour with a lightness value of 80.46±0.01; yield of 23.50%; viscosity 99.33 cP, gel strength 307.63 g/cm2, sulfate 13.29% (w/w), ash 8.33% (w/w) and acid-insoluble ash 1.33% (w/w).. |
| 67. |
Mari Takahara, Rie Wakabayashi, Naoki Fujimoto, Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Enzymatic Cell-Surface Decoration with Proteins using Amphiphilic Lipid-Fused Peptide Substrates, Chemistry - A European Journal, 10.1002/chem.201900370, 25, 30, 7315-7321, 2019.05, © 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim Lipid modification of proteins plays a significant role in the activation of cellular signals such as proliferation. Thus, the demand for lipidated proteins is rising. However, getting a high yield and purity of lipidated proteins has been challenging. We developed a strategy for modifying proteins with a wide variety of synthetic lipids using microbial transglutaminase (MTG), which catalyzes the cross-linking reaction between a specific glutamine (Q) in a protein and lysine (K) in the lipid-fused peptide. The synthesized lipid-G3S-MRHKGS lipid (lipid: fatty acids, tocopherol, lithocholic acid, cholesterol) was successfully conjugated to a protein fused with LLQG (Q-tagged protein) by an MTG reaction, yielding >90 % conversion of the Q-tagged protein in a lipidated form. The purified lipid–protein conjugates were used for labeling the cell membrane in vitro, resulting in best-anchoring ability of cholesterol modification. Furthermore, in situ cell-surface decoration with the protein was established in a simple manner: subjection of cells to a mixture of cholesterol-fused peptides, Q-tagged proteins and MTG.. |
| 68. |
Functional horseradish peroxidase-streptavidin chimeric proteins prepared using a silkworm-baculovirus expression system for diagnostic purposes.. |
| 69. |
In vivo biocompatibility, pharmacokinetics, antitumor efficacy, and hypersensitivity evaluation of ionic liquid-mediated paclitaxel formulations. |
| 70. |
Djohan Yovita, Azukizawa Tomoki, Patmawati, Sakai Kotaro, Yano Yuki, Sato Fumiya, Takahashi Ryoji, Yohda Masafumi, Maeda Mizuo, Kamiya Noriho, Zako Tamotsu, Molecular chaperone prefoldin-assisted biosynthesis of gold nanoparticles with improved size distribution and dispersion, BIOMATERIALS SCIENCE, 10.1039/c8bm01026a, 7, 5, 1801-1804, 2019.05. |
| 71. |
Synthesis and characterization of choline–fatty-acid-based ionic liquids: A new biocompatible surfactant. |
| 72. |
Transcutaneous Codelivery of Tumor Antigen and Resiquimod in Solid-in-Oil Nanodispersions Promotes Antitumor Immunity. |
| 73. |
昆虫工場を利用したウイルス様粒子ワクチン生産系の確立とその応用. |
| 74. |
Muhamad Alif Razi, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya, Self-Assembled Reduced Albumin and Glycol Chitosan Nanoparticles for Paclitaxel Delivery, Langmuir, 10.1021/acs.langmuir.8b02809, 35, 7, 2610-2618, 2019.02, Cancer continues to pose health problems for people all over the world. Nanoparticles (NPs) have emerged as a promising platform for effective cancer chemotherapy. NPs formed by the assembly of proteins and chitosan (CH) through noncovalent interactions are attracting a great deal of interest. However, the poor water solubility of CH and low stability of this kind of NP limit its practical application. Herein, the formation of reduced bovine serum albumin (rBSA) and glycol chitosan (GC) nanoparticles (rBG-NPs) stabilized by hydrophobic interactions and disulfide bonds was demonstrated for paclitaxel (PTX) delivery. The effects of the rBSA:GC mass ratio and pH on the particle size, polydispersity index (PDI), number of particles, and surface charge were evaluated. The formation mechanism and stability of the NPs were determined by compositional analysis and dynamic light scattering. Hydrophobic and electrostatic interactions were the driving forces for the formation of the rBG-NPs, and the NPs were stable under physiological conditions. PTX was successfully encapsulated into rBG-NPs with a high encapsulation efficiency (90%). PTX-loaded rBG-NPs had a particle size of 400 nm with a low PDI (0.2) and positive charge. rBG-NPs could be internalized by HeLa cells, possibly via endocytosis. An in vitro cytotoxicity study revealed that PTX-loaded rBG-NPs had anticancer activity that was lower than that of a Taxol-like formulation at 24 h but had similar activity at 48 h, possibly because of the slow release of PTX into the cells. Our study suggests that rBG-NPs could be used as a potential nanocarrier for hydrophobic drugs.. |
| 75. |
Self-Assembled Reduced Albumin and Glycol Chitosan Nanoparticles for Paclitaxel Delivery. |
| 76. |
Rie Wakabayashi, Ayumi Suehiro, Masahiro Goto, Noriho Kamiya, Designer aromatic peptide amphiphiles for self-assembly and enzymatic display of proteins with morphology control, Chemical Communications, 10.1039/C8CC08163H, 55, 5, 640-643, 2019.01, We herein designed bi-functional aromatic peptide amphiphiles both self-assembling to fibrous nanomaterials and working as a substrate of microbial transglutaminase, leading to peptidyl scaffolds with different morphologies that can be enzymatically post-functionalized with proteins.. |
| 77. |
Shuto Kozaka, Rie Wakabayashi, Onofrio Annunziata, Milan Balaz, Masahiro Goto, Noriho Kamiya, Sergei V. Dzyuba, Effect of macromolecular crowding on the conformational behaviour of a porphyrin rotor, Journal of Photochemistry and Photobiology A: Chemistry, 10.1016/j.jphotochem.2018.10.006, 369, 115-118, 2019.01, Macromolecular crowding modulates the conformational preference of a small molecular rotor, which is used as a molecular viscometer. The shift towards the planar conformation of the porphyrin rotor was observed in the presence of increasing concentrations and sizes of polyethylene glycols. This observation highlights the differences between the behaviour of this molecular viscometer in the crowding and non-crowding types of media.. |
| 78. |
Safrina Dyah Hardiningtyas, Rie Wakabayashi, Ryutaro Ishiyama, Yuki Owada, Masahiro Goto, Noriho Kamiya, Enhanced potential of therapeutic applications of curcumin using solid-in-water nanodispersion technique, JOURNAL OF CHEMICAL ENGINEERING OF JAPAN, 10.1252/jcej.18we060, 52, 1, 138-143, 2019.01, Curcumin (Cur), a hydrophobic polyphenol compound, holds promising potential as an anticancer agent. However, the poor solubility in water and the low bioavailability of curcumin have limited its therapeutic applications. In this regard, we reported the formulation of curcumin using a solid-in-water (S/W) nanodispersion technique to enhance the water solubility and therapeutic activity of curcumin. This new aqueous formulation comprises simple preparation protocols: emulsification and freeze-drying for encapsulating hydrophobic biomolecules with a hydrophilic surfactant, followed by redispersion of the resultant solid complexes in an aqueous solution. Pluronics F68 and F127 were used here for the encapsulation of curcumin. Enhanced aqueous solubility of curcumin was achieved by encapsulating curcumin with a hydrophilic surfactant using the S/W nanodispersion technique. The resultant nanosized formulation had a narrow size distribution and high entrapment efficiency of curcumin. The highest loading capacity of curcumin in S/W nanodispersion was obtained with a weight ratio of curcumin to pluronic of 1: 10 for both surfactants. The release profile of the complexes was found to depend on the type of surfactant, suggesting that the selection of a proper surfactant is crucial for controlling curcumin delivery. The anticancer activity of the S/W formulation of curcumin was correlated with the drug release profiles and cellular uptake, which in turn was influenced by the hydrophobicity and chemical structure of the surfactant.. |
| 79. |
Dani Permana, Kosuke Minamihata, Tsuneyuki Tatsuke, Jae M. Lee, Takahiro Kusakabe, Masahiro Goto, Noriho Kamiya, Polymerization of Horseradish Peroxidase by a Laccase-Catalyzed Tyrosine Coupling Reaction, Biotechnology Journal, 10.1002/biot.201800531, 2019.01, The polymerization of proteins can create newly active and large bio-macromolecular assemblies that exhibit unique functionalities depending on the properties of the building block proteins and the protein units in polymers. Herein, the first enzymatic polymerization of horseradish peroxidase (HRP) is reported. Recombinant HRPs fused with a tyrosine-tag (Y-tag) through a flexible linker at the N- and/or C-termini are expressed in silkworm, Bombyx mori. Trametes sp. laccase (TL) is used to activate the tyrosine of Y-tagged HRPs with molecular O
2
to form a tyrosyl-free radical, which initiates the tyrosine coupling reaction between the HRP units. A covalent dityrosine linkage is also formed through a HRP-catalyzed self-crosslinking reaction in the presence of H
2
O
2
. The addition of H
2
O
2
in the self-polymerization of Y-tagged HRPs results in lower activity of the HRP polymers, whereas TL provides site-selectivity, mild reaction conditions and maintains the activity of the polymeric products. The cocrosslinking of Y-tagged HRPs and HRP-protein G (Y-HRP-pG) units catalyzed by TL shows a higher signal in enzyme-linked immunosorbent assay (ELISA) than the genetically pG-fused HRP, Y-HRP-pG, and its polymers. This new enzymatic polymerization of HRP promises to provide highly active and functionalized polymers for biomedical applications and diagnostics probes.. |
| 80. |
Rie Wakabayashi, Hiroki Obayashi, Ryuichiro Hashimoto, Noriho Kamiya, Masahiro Goto, Complementary interaction with peptide amphiphiles guides size-controlled assembly of small molecules for intracellular delivery, Chemical Communications, 10.1039/c9cc02473e, 55, 49, 6997-7000, 2019.01, We introduced complementary interactions between peptide amphiphiles and a small fluorescence dye to develop a programmable multi-component supramolecular assembly, and intracellular delivery of the dye was controlled by the dimensions of the co-assembly, which was manipulated by the peptide design.. |
| 81. |
Md Raihan Chowdhury, Rahman Md Moshikur, Rie Wakabayashi, Yoshiro Tahara, Noriho Kamiya, Muhammad Moniruzzaman, Masahiro Goto, Development of a novel ionic liquid-curcumin complex to enhance its solubility, stability, and activity, Chemical Communications, 10.1039/c9cc02812a, 55, 54, 7737-7740, 2019.01, We report a one-step emulsification and rapid freeze-drying process to develop a curcumin-ionic liquid (CCM-IL) complex that could be readily dispersed in water with a significantly enhanced solubility of ∼8 mg mL-1 and half-life (t1/2) of ∼260 min compared with free CCM (solubility ∼30 nM and t1/2 ∼ 20 min). This process using an IL consisting of a long chain carbon backbone as a surfactant, may provide an alternative way of enhancing the solubility of poorly water-soluble drugs.. |
| 82. |
Rie Wakabayashi, Hidetoshi Kono, Shuto Kozaka, Yoshiro Tahara, Noriho Kamiya, Masahiro Goto, Transcutaneous codelivery of tumor antigen and resiquimod in solid-in-oil nanodispersions promotes antitumor immunity, ACS Biomaterials Science and Engineering, 10.1021/acsbiomaterials.9b00260, 2019.01, Cancer vaccines aim to prevent or inhibit tumor growth by inducing an immune response to tumor-associated antigens (TAAs) encoded by or present in the vaccine. Previous work has demonstrated that effective antitumor immunity can be induced using a codelivery system in which nonspecific immunostimulatory molecules are administered together with TAAs. In this study, we investigated the antitumor effects of a solid-in-oil (S/O) nanodispersion system containing a model TAA, ovalbumin (OVA), and resiquimod (R-848), a small molecular Toll-like receptor 7/8 ligand, which induces an antigen-nonspecific cellular immune response that is crucial for the efficacy of cancer vaccines. R-848 was contained in the outer oil phase of S/O nanodispersion. Analysis of OVA and R-848 permeation in mouse skin after application of an R-848 S/O nanodispersion indicated that R-848 rapidly permeated the skin and preactivated Langerhans cells, resulting in efficient uptake of OVA and migration of antigen-loaded Langerhans cells to the draining lymph nodes. Transcutaneous immunization of mice with an R-848 S/O nanodispersion inhibited the growth of E.G7-OVA tumors and prolonged mouse survival to a greater extent than did immunization with an S/O nanodispersion containing OVA alone. Consistent with this observation, antigen-specific secretion of the Th1 cytokine interferon-γand cytolytic activity were both high in splenocytes isolated from mice immunized with R-848 S/O. Our results thus demonstrate that codelivery of R-848 significantly amplified the antitumor immune response induced by antigen-containing S/O nanodispersions and further suggest that S/O nanodispersions may be effective formulations for codelivery of TAAs and R-848 in transcutaneous cancer vaccines.. |
| 83. |
Effect of macromolecular crowding on the conformational behaviour of a porphyrin rotor. |
| 84. |
Enhanced Potential of Therapeutic Applications of Curcumin Using Solid-in-Water Nanodispersion Technique. |
| 85. |
Noriho Kamiya, Yuki Ohama, Kosuke Minamihata, Rie Wakabayashi, Masahiro Goto, Liquid Marbles as an Easy-to-Handle Compartment for Cell-Free Synthesis and In Situ Immobilization of Recombinant Proteins, Biotechnology Journal, 10.1002/biot.201800085, 13, 12, 2018.12, Liquid marble (LM), a self-standing micro-scale aqueous droplet, emerges as a micro-bioreactor in biological applications. Herein, the potential of LM as media for cell-free synthesis and simultaneous immobilization of recombinant proteins is explored. Initially, formation of hydrogel marble (HM) by using an enzymatic disulfide-based hydrogelation technique is confirmed by incorporating three components, horseradish peroxidase (HRP), a tetra-thiolated poly(ethylene glycol) derivative, and glycyl-L-tyrosine, in LM. The compatibility of the enzymatic hydrogelation with cell-free protein synthesis in LM is then validated. Although the hydrogelation reduces the level of protein synthesis in LM when compared with that in a test tube, the biosynthesis of enhanced green fluorescent protein (EGFP) is achieved. Interestingly, EGFP synthesized in LM is entrapped in the HM, and the introduction of a cysteine residue to EGFP by genetic engineering further increases the amount of protein immobilization in the hydrogel matrices. These results suggest that the cell-free synthesis and HRP-catalyzed hydrogelation can be conducted in parallel in LM, and the eventual entrapment of the key components in HM is possible. Facile recovery of macromolecular products immobilized in HM by degrading the hydrogel network under reducing conditions should lead to the design of an easy-to-handle system to screen protein functions.. |
| 86. |
Designer aromatic peptide amphiphiles for self-assembly and enzymatic display of proteins with morphology control. |
| 87. |
Noriho Kamiya, Yuki Ohama, Kosuke Minamihata, Rie Wakabayashi, Masahiro Goto, Liquid Marbles as an Easy-to-Handle Compartment for Cell-Free Synthesis and In Situ Immobilization of Recombinant Proteins., Biotechnology journal, 10.1002/biot.201800085, 13, 12, e1800085, 2018.12, Liquid marble (LM), a self-standing micro-scale aqueous droplet, emerges as a micro-bioreactor in biological applications. Herein, the potential of LM as media for cell-free synthesis and simultaneous immobilization of recombinant proteins is explored. Initially, formation of hydrogel marble (HM) by using an enzymatic disulfide-based hydrogelation technique is confirmed by incorporating three components, horseradish peroxidase (HRP), a tetra-thiolated poly(ethylene glycol) derivative, and glycyl-L-tyrosine, in LM. The compatibility of the enzymatic hydrogelation with cell-free protein synthesis in LM is then validated. Although the hydrogelation reduces the level of protein synthesis in LM when compared with that in a test tube, the biosynthesis of enhanced green fluorescent protein (EGFP) is achieved. Interestingly, EGFP synthesized in LM is entrapped in the HM, and the introduction of a cysteine residue to EGFP by genetic engineering further increases the amount of protein immobilization in the hydrogel matrices. These results suggest that the cell-free synthesis and HRP-catalyzed hydrogelation can be conducted in parallel in LM, and the eventual entrapment of the key components in HM is possible. Facile recovery of macromolecular products immobilized in HM by degrading the hydrogel network under reducing conditions should lead to the design of an easy-to-handle system to screen protein functions.. |
| 88. |
Dani Permana, Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Laccase-catalyzed bioconjugation of tyrosine-tagged functional proteins, Journal of Bioscience and Bioengineering, 10.1016/j.jbiosc.2018.05.013, 126, 5, 559-566, 2018.11, The site-specific cross-linking of functional proteins creates macromolecular assemblies that exhibit unique biochemical and/or physicochemical properties. Herein, we explored the potential of laccase as a biocatalyst for the site-specific cross-linking of tyrosine-tagged proteins. Trametes sp. laccase (TL) was selected as the cross-linking catalyst, and Escherichia coli alkaline phosphatase (BAP) and antibody-binding proteins (pG2pAs) were employed as model proteins. The protein models were genetically fused to a peptide tag containing a tyrosine residue (Y-tag) at the N- and/or C-termini. Proteins without Y-tags were used as controls. The Y-tagged proteins could be recognized by TL as macromolecular substrates, leading to the oxidative formation of protein polymers, whereas no polymerization was observed with intact BAP or pG2pA. The TL-catalyzed cross-linking of Y-tagged proteins proceeded at a relatively high pH in comparison with that of small phenolic substrates. Co-polymers of BAP and pG2pA were able to be prepared by mixing the aqueous solution of each component in the presence of TL. A combination of bis-Y-tagged pG2pA (Y-pG2pA-Y) and Y-tagged BAP (BAP-Y) yielded functional co-polymers compatible with enzyme-linked immunosorbent assay (ELISA). The detection limit of the ELISA of ovalbumin with anti-OVA IgG depended on the molar ratio of BAP-Y and Y-pG2pA-Y in the TL-catalyzed cross-linking reaction. A high molar ratio of BAP-Y to Y-pG2pA-Y (75:1) resulted in the highest absorbance in the ELISA. The results suggested that the formation of a bifunctional protein polymer with a high molar ratio of signaling unit to antibody-binding unit gave better performance in antigen detection than using lower ratios.. |
| 89. |
Dani Permana, Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Laccase-catalyzed bioconjugation of tyrosine-tagged functional proteins., Journal of bioscience and bioengineering, 10.1016/j.jbiosc.2018.05.013, 126, 5, 559-566, 2018.11, The site-specific cross-linking of functional proteins creates macromolecular assemblies that exhibit unique biochemical and/or physicochemical properties. Herein, we explored the potential of laccase as a biocatalyst for the site-specific cross-linking of tyrosine-tagged proteins. Trametes sp. laccase (TL) was selected as the cross-linking catalyst, and Escherichia coli alkaline phosphatase (BAP) and antibody-binding proteins (pG2pAs) were employed as model proteins. The protein models were genetically fused to a peptide tag containing a tyrosine residue (Y-tag) at the N- and/or C-termini. Proteins without Y-tags were used as controls. The Y-tagged proteins could be recognized by TL as macromolecular substrates, leading to the oxidative formation of protein polymers, whereas no polymerization was observed with intact BAP or pG2pA. The TL-catalyzed cross-linking of Y-tagged proteins proceeded at a relatively high pH in comparison with that of small phenolic substrates. Co-polymers of BAP and pG2pA were able to be prepared by mixing the aqueous solution of each component in the presence of TL. A combination of bis-Y-tagged pG2pA (Y-pG2pA-Y) and Y-tagged BAP (BAP-Y) yielded functional co-polymers compatible with enzyme-linked immunosorbent assay (ELISA). The detection limit of the ELISA of ovalbumin with anti-OVA IgG depended on the molar ratio of BAP-Y and Y-pG2pA-Y in the TL-catalyzed cross-linking reaction. A high molar ratio of BAP-Y to Y-pG2pA-Y (75:1) resulted in the highest absorbance in the ELISA. The results suggested that the formation of a bifunctional protein polymer with a high molar ratio of signaling unit to antibody-binding unit gave better performance in antigen detection than using lower ratios.. |
| 90. |
Mari Takahara, Rie Wakabayashi, Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Design of Lipid-Protein Conjugates Using Amphiphilic Peptide Substrates of Microbial Transglutaminase, ACS App. Bio Mater., 10.1021/acsabm.8b00271, 1, 6, 1823-1829, 2018.10, Lipid modification of proteins plays a significant role in the activation of cellular signals such as proliferation. Thus, the demand for lipidated proteins is rising. However, getting a high yield and purity of lipidated proteins has been challenging. We developed a strategy for modifying proteins with a wide variety of synthetic lipids using microbial transglutaminase (MTG), which catalyzes the cross-linking reaction between a specific glutamine (Q) in a protein and lysine (K) in the lipid-fused peptide. The synthesized lipid-G3S-MRHKGS lipid (lipid: fatty acids, tocopherol, lithocholic acid, cholesterol) was successfully conjugated to a protein fused with LLQG (Q-tagged protein) by an MTG reaction, yielding >90 % conversion of the Q-tagged protein in a lipidated form. The purified lipid–protein conjugates were used for labeling the cell membrane in vitro, resulting in best-anchoring ability of cholesterol modification. Furthermore, in situ cell-surface decoration with the protein was established in a simple manner: subjection of cells to a mixture of cholesterol-fused peptides, Q-tagged proteins and MTG.. |
| 91. |
Design of lipid-protein conjugate with a self-assembling ability on a cell membrane by using microbial transglutaminase reaction. |
| 92. |
Safrina Dyah Hardiningtyas, Rie Wakabayashi, Momoko Kitaoka, Yoshiro Tahara, Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Mechanistic investigation of transcutaneous protein delivery using solid-in-oil nanodispersion
A case study with phycocyanin, European Journal of Pharmaceutics and Biopharmaceutics, 10.1016/j.ejpb.2018.01.020, 127, 44-50, 2018.06, Phycocyanin (PC), a water-soluble protein-chromophore complex composed of hexameric (αβ)6 subunits, has important biological functions in blue-green algae as well as pharmacological activities in biomedicine. We have previously developed a solid-in-oil (S/O) nanodispersion method to deliver biomacromolecules through the skin, although the transcutaneous mechanism has not yet been fully elucidated. To study the mechanism of transcutaneous protein delivery, we therefore enabled S/O nanodispersion by coating PC with hydrophobic surfactants and evaluated how the proteinaceous macromolecules formulated in an oil phase might permeate the skin. The extent of S/O nanodispersion of PC was dependent on the type of surfactant, suggesting that the selection of a suitable surfactant is crucial for encapsulating a large protein having a subunit structure. By measuring the intrinsic fluorescence of PC, we found that S/O nanodispersion facilitated the accumulation of PC in the stratum corneum (SC) of Yucatan micropig skin. Furthermore, after crossing the SC layer, the fluorescent recovery of PC was evident, indicating the release of the biologically active form of PC from the SC into the deeper skin layer.. |
| 93. |
Akitsu Masuda, Jian Xu, Kosuke Minamihata, Genki Kagawa, Yusei Hamada, Yoshiki Morifuji, Takumi Yano, Masato Hino, Daisuke Morokuma, Noriko Karasaki, Hiroaki Mon, Noriho Kamiya, Takahiro Kusakabe, Jae Man Lee, Production of a biologically active human basic fibroblast growth factor using silkworm-baculovirus expression vector system, Journal of Asia-Pacific Entomology, 10.1016/j.aspen.2018.05.002, 21, 2, 716-720, 2018.06, As a therapeutic treatment, recombinant human basic fibroblast growth factor (rhbFGF) is usually employed in tissue regeneration, and as an essential component in culture medium for maintaining the induced pluripotent stem (iPS) cell and embryonic stem (ES) cell in an undifferentiated state. Therefore, a large amount of biologically active rhbFGF is required. In this study, silkworm-baculovirus expression vector system (silkworm-BEVS) is employed to achieve a high productivity of recombinant rhbFGF with two small affinity tags (His-tag and STREP-tag) at the N or C-terminus. It is observed that rhbFGF with 30 K signal peptide of silkworm were successfully expressed but are not sufficiently secreted into the culture medium of cultured insect cells. Then we purified the N- or C-tagged intracellular rhbFGF protein and obtained a yield of about 0.7 mg/larva and 1.2 mg/larva, respectively. Although the final yield of the C-tagged rhbFGF is higher than that of the N-tagged, rhbFGF with N-tag demonstrated promising and comparable biological activity, which is evaluated through a mammalian cell proliferation assay. Taken together, these results indicate that silkworm-BEVS could contribute to the mass-production of the biologically active rhbFGF for medical uses.. |
| 94. |
Patma, Kosuke Minamihata, T. Tatsuke, Jae Man Lee, Takahiro Kusakabe, Noriho Kamiya, Expression and Activation of Horseradish Peroxidase–Protein A/G Fusion Protein in Silkworm Larvae for Diagnostic Purposes, Biotechnology Journal, 10.1002/biot.201700624, 13, 1700624, 2018.06. |
| 95. |
Md Raihan Chowdhury, Rahman Md Moshikur, Rie Wakabayashi, Yoshiro Tahara, Noriho Kamiya, Muhammad Moniruzzaman, Masahiro Goto, Ionic-Liquid-Based Paclitaxel Preparation
A New Potential Formulation for Cancer Treatment, Molecular pharmaceutics, 10.1021/acs.molpharmaceut.8b00305, 15, 6, 2484-2488, 2018.06, Paclitaxel (PTX) injection (i.e., Taxol) has been used as an effective chemotherapeutic treatment for various cancers. However, the current Taxol formulation contains Cremophor EL, which causes hypersensitivity reactions during intravenous administration and precipitation by aqueous dilution. This communication reports the preliminary results on the ionic liquid (IL)-based PTX formulations developed to address the aforementioned issues. The formulations were composed of PTX/cholinium amino acid ILs/ethanol/Tween-80/water. A significant enhancement in the solubility of PTX was observed with considerable correlation with the density and viscosity of the ILs, and with the side chain of the amino acids used as anions in the ILs. Moreover, the formulations were stable for up to 3 months. The driving force for the stability of the formulation was hypothesized to be the involvement of different types of interactions between the IL and PTX. In vitro cytotoxicity and antitumor activity of the IL-based formulations were evaluated on HeLa cells. The IL vehicles without PTX were found to be less cytotoxic than Taxol, while both the IL-based PTX formulation and Taxol exhibited similar antitumor activity. Finally, in vitro hypersensitivity reactions were evaluated on THP-1 cells and found to be significantly lower with the IL-based formulation than Taxol. This study demonstrated that specially designed ILs could provide a potentially safer alternative to Cremophor EL as an effective PTX formulation for cancer treatment giving fewer hypersensitivity reactions.. |
| 96. |
Patmawati Xxxx, Kosuke Minamihata, Tsuneyuki Tatsuke, Jae Man Lee, Takahiro Kusakabe, Noriho Kamiya, Expression and Activation of Horseradish Peroxidase-Protein A/G Fusion Protein in Silkworm Larvae for Diagnostic Purposes., Biotechnology journal, 10.1002/biot.201700624, 13, 6, e1700624, 2018.06, Recombinant protein production can create artificial proteins with desired functions by introducing genetic modifications to the target proteins. Horseradish peroxidase (HRP) has been used extensively as a reporter enzyme in biotechnological applications; however, recombinant production of HRP has not been very successful, hampering the utilization of HRP with genetic modifications. A fusion protein comprising an antibody binding protein and HRP will be an ideal bio-probe for high-quality HRP-based diagnostic systems. A HRP-protein A/G fusion protein (HRP-pAG) is designed and its production in silkworm (Bombyx mori) is evaluated for the first time. HRP-pAG is expressed in a soluble apo form, and is activated successfully by incubating with hemin. The activated HRP-pAG is used directly for ELISA experiments and retains its activity over 20 days at 4 °C. Moreover, HRP-pAG is modified with biotin by the microbial transglutaminase (MTG) reaction. The biotinylated HRP-pAG is conjugated with streptavidin to form a HRP-pAG multimer and the multimeric HRP-pAG produced higher signals in the ELISA system than monomeric HRP-pAG. The successful production of recombinant HRP in silkworm will contribute to creating novel HRP-based bioconjugates as well as further functionalization of HRP by applying enzymatic post-translational modifications.. |
| 97. |
Md. Raihan Chowdhury, Rahman Md Moshikur, Rie Wakabayashi, Yoshiro Tahara, Noriho Kamiya, Muhammad Moniruzzaman, Masahiro Goto, Ionic-Liquid-Based Paclitaxel Preparation: A New Potential Formulation for Cancer Treatment, Molecular Pharmaceutics, 10.1021/acs.molpharmaceut.8b00305, 15, 6, 2484-2488, 2018.06, Paclitaxel (PTX) injection (i.e., Taxol) has been used as an effective chemotherapeutic treatment for various cancers. However, the current Taxol formulation contains Cremophor EL, which causes hypersensitivity reactions during intravenous administration and precipitation by aqueous dilution. This communication reports the preliminary results on the ionic liquid (IL)-based PTX formulations developed to address the aforementioned issues. The formulations were composed of PTX/cholinium amino acid ILs/ethanol/Tween-80/water. A significant enhancement in the solubility of PTX was observed with considerable correlation with the density and viscosity of the ILs, and with the side chain of the amino acids used as anions in the ILs. Moreover, the formulations were stable for up to 3 months. The driving force for the stability of the formulation was hypothesized to be the involvement of different types of interactions between the IL and PTX. In vitro cytotoxicity and antitumor activity of the IL-based formulations were evaluated on HeLa cells. The IL vehicles without PTX were found to be less cytotoxic than Taxol, while both the IL-based PTX formulation and Taxol exhibited similar antitumor activity. Finally, in vitro hypersensitivity reactions were evaluated on THP-1 cells and found to be significantly lower with the IL-based formulation than Taxol. This study demonstrated that specially designed ILs could provide a potentially safer alternative to Cremophor EL as an effective PTX formulation for cancer treatment giving fewer hypersensitivity reactions.. |
| 98. |
Safrina Dyah Hardiningtyas, Rie Wakabayashi, Momoko Kitaoka, Yoshiro Tahara, Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Mechanistic investigation of transcutaneous protein delivery using solid-in-oil nanodispersion: A case study with phycocyanin., European journal of pharmaceutics and biopharmaceutics : official journal of Arbeitsgemeinschaft fur Pharmazeutische Verfahrenstechnik e.V, 10.1016/j.ejpb.2018.01.020, 127, 44-50, 2018.06, Phycocyanin (PC), a water-soluble protein-chromophore complex composed of hexameric (αβ)6 subunits, has important biological functions in blue-green algae as well as pharmacological activities in biomedicine. We have previously developed a solid-in-oil (S/O) nanodispersion method to deliver biomacromolecules through the skin, although the transcutaneous mechanism has not yet been fully elucidated. To study the mechanism of transcutaneous protein delivery, we therefore enabled S/O nanodispersion by coating PC with hydrophobic surfactants and evaluated how the proteinaceous macromolecules formulated in an oil phase might permeate the skin. The extent of S/O nanodispersion of PC was dependent on the type of surfactant, suggesting that the selection of a suitable surfactant is crucial for encapsulating a large protein having a subunit structure. By measuring the intrinsic fluorescence of PC, we found that S/O nanodispersion facilitated the accumulation of PC in the stratum corneum (SC) of Yucatan micropig skin. Furthermore, after crossing the SC layer, the fluorescent recovery of PC was evident, indicating the release of the biologically active form of PC from the SC into the deeper skin layer.. |
| 99. |
Muhamad Alif Razi, Rie Wakabayashi, Yoshiro Tahara, Masahiro Goto, Noriho Kamiya, Genipin-stabilized caseinatehitosan nanoparticles for enhanced stability and anti-cancer activity of curcumin, Colloids and Surfaces B: Biointerfaces, 10.1016/j.colsurfb.2018.01.041, 164, 308-315, 2018.04, Nanoparticles formed by the assembly of protein and polysaccharides are of great interest for the delivery of hydrophobic molecules. Herein, the formation of genipin-crosslinked nanoparticles from caseinate (CS) and chitosan (CH) is reported for the delivery of curcumin, a polyphenolic compound from turmeric, to cells. Genipin-crosslinked CS-CH nanoparticles (G-CCNPs) having a diameter of ∼250 nm and a low polydispersity index showed excellent stability over a wide pH range, as indicated by dynamic light scattering and transmission electron microscopic measurements. Cellular uptake of curcumin loaded into G-CCNPs by HeLa cells was improved, as measured by confocal laser scanning microscopy (CLSM) and fluorescence-activated cell-sorting analysis. Cell proliferation assays indicated that G-CCNPs were nontoxic and that curcumin's anticancer activity in vitro was also improved by G-CCNPs. Stability of curcumin at neutral pH was enhanced by G-CCNPs. CLSM study revealed that G-CCNPs were poorly internalized by HeLa cells, possibly because of strong cell membrane interactions and a negative zeta potential. Overall, our results suggested that the enhanced curcumin cytotoxicity might be associated with the enhanced stability of curcumin by G-CCNPs and free curcumin released from G-CCNPs into the cell. These biocompatible NPs might be suitable carriers for enhancing curcumin's therapeutic potential.. |
| 100. |
Muhamad Alif Razi, Rie Wakabayashi, Yoshiro Tahara, Masahiro Goto, Noriho Kamiya, Genipin-stabilized caseinatehitosan nanoparticles for enhanced stability and anti-cancer activity of curcumin, Colloids and Surfaces B: Biointerfaces, 10.1016/j.colsurfb.2018.01.041, 164, 308-315, 2018.04, Nanoparticles formed by the assembly of protein and polysaccharides are of great interest for the delivery of hydrophobic molecules. Herein, the formation of genipin-crosslinked nanoparticles from caseinate (CS) and chitosan (CH) is reported for the delivery of curcumin, a polyphenolic compound from turmeric, to cells. Genipin-crosslinked CS-CH nanoparticles (G-CCNPs) having a diameter of ∼250 nm and a low polydispersity index showed excellent stability over a wide pH range, as indicated by dynamic light scattering and transmission electron microscopic measurements. Cellular uptake of curcumin loaded into G-CCNPs by HeLa cells was improved, as measured by confocal laser scanning microscopy (CLSM) and fluorescence-activated cell-sorting analysis. Cell proliferation assays indicated that G-CCNPs were nontoxic and that curcumin's anticancer activity in vitro was also improved by G-CCNPs. Stability of curcumin at neutral pH was enhanced by G-CCNPs. CLSM study revealed that G-CCNPs were poorly internalized by HeLa cells, possibly because of strong cell membrane interactions and a negative zeta potential. Overall, our results suggested that the enhanced curcumin cytotoxicity might be associated with the enhanced stability of curcumin by G-CCNPs and free curcumin released from G-CCNPs into the cell. These biocompatible NPs might be suitable carriers for enhancing curcumin's therapeutic potential.. |
| 101. |
Geisa A.L.G. Budinova, Yutaro Mori, Tsutomu Tanaka, Noriho Kamiya, Casein-based scaffold for artificial cellulosome design, Process Biochemistry, 10.1016/j.procbio.2017.12.013, 66, 140-145, 2018.03, Cellulosomal systems are known as highly efficient biocatalysts in the degradation of lignocellulosic biomass in nature, but they remain unsuitable for industrial applications. In seeking alternatives to natural cellulosomes, casein was chosen as a scaffold for cellulase clustering. Casein is recognized as an excellent substrate for microbial transglutaminase (MTG) because it contains naturally reactive glutamine and lysine residues. A substrate peptide containing an MTG-reactive lysine residue was inserted into the C-terminus of the endoglucanase Cel5A and Cel6A from Thermobifida fusca using genetic engineering. The engineered cellulases, EG(Cel5A) and EG(Cel6A), were conjugated onto casein in different ratios by an MTG-mediated site-specific protein crosslinking reaction. Overall, a more than two-fold increase was observed when EG(Cel5A) was conjugated onto N,N-dimethylcasein, but a small or no change was observed for EG(Cel6A).. |
| 102. |
Rie Wakabayashi, Masato Sakuragi, Shuto Kozaka, Yoshiro Tahara, Noriho Kamiya, Masahiro Goto, Solid-in-Oil Peptide Nanocarriers for Transcutaneous Cancer Vaccine Delivery against Melanoma, Molecular Pharmaceutics, 10.1021/acs.molpharmaceut.7b00894, 15, 3, 955-961, 2018.03, Cancer vaccines represent a prophylactic or therapeutic method of suppressing cancer by activating the adaptive immune system. The immune response is initiated by the delivery of tumor antigens to antigen presenting cells (APCs). The use of peptides as vaccine antigens is advantageous, especially in the availability and productivity of pure and defined antigens. However, their limited immunogenicity remains a major drawback, and therefore, the utilization of nanocarriers as a means of delivering antigens to target cells and/or the addition of immune stimulants have been investigated as an efficient peptide-based cancer vaccine. We have developed a solid-in-oil (S/O) nanodispersion as a transcutaneous nanocarrier for hydrophilic molecules. This system has attractive features as a peptide nanocarrier for cancer vaccines, including transcutaneous targeting of professional APCs in the skin, high encapsulation efficacy of hydrophilic molecules, and capacity for coloading with a variety of immune stimulants such as adjuvants. We therefore sought to utilize the developed S/O nanodispersion for the delivery of the tyrosine-related protein 2 peptide, TRP-2180-188, as a peptide antigen against melanoma. Transcutaneous vaccination of the S/O nanodispersion coloaded with adjuvant R-848 was associated with a significant inhibition of melanoma growth and suppression of lung metastasis in tumor-bearing mice. Our findings indicate the potential of S/O nanodispersions as an endogenous peptide carrier for cancer vaccines.. |
| 103. |
Geisa A.L.G. Budinova, Yutaro Mori, Tsutomu Tanaka, Noriho Kamiya, Casein-based scaffold for artificial cellulosome design, Process Biochemistry, 10.1016/j.procbio.2017.12.013, 66, 140-145, 2018.03, Cellulosomal systems are known as highly efficient biocatalysts in the degradation of lignocellulosic biomass in nature, but they remain unsuitable for industrial applications. In seeking alternatives to natural cellulosomes, casein was chosen as a scaffold for cellulase clustering. Casein is recognized as an excellent substrate for microbial transglutaminase (MTG) because it contains naturally reactive glutamine and lysine residues. A substrate peptide containing an MTG-reactive lysine residue was inserted into the C-terminus of the endoglucanase Cel5A and Cel6A from Thermobifida fusca using genetic engineering. The engineered cellulases, EG(Cel5A) and EG(Cel6A), were conjugated onto casein in different ratios by an MTG-mediated site-specific protein crosslinking reaction. Overall, a more than two-fold increase was observed when EG(Cel5A) was conjugated onto N,N-dimethylcasein, but a small or no change was observed for EG(Cel6A).. |
| 104. |
Rie Wakabayashi, Masato Sakuragi, Shuto Kozaka, Yoshiro Tahara, Noriho Kamiya, Masahiro Goto, Solid-in-Oil Peptide Nanocarriers for Transcutaneous Cancer Vaccine Delivery against Melanoma, Molecular Pharmaceutics, 10.1021/acs.molpharmaceut.7b00894, 15, 3, 955-961, 2018.03, Cancer vaccines represent a prophylactic or therapeutic method of suppressing cancer by activating the adaptive immune system. The immune response is initiated by the delivery of tumor antigens to antigen presenting cells (APCs). The use of peptides as vaccine antigens is advantageous, especially in the availability and productivity of pure and defined antigens. However, their limited immunogenicity remains a major drawback, and therefore, the utilization of nanocarriers as a means of delivering antigens to target cells and/or the addition of immune stimulants have been investigated as an efficient peptide-based cancer vaccine. We have developed a solid-in-oil (S/O) nanodispersion as a transcutaneous nanocarrier for hydrophilic molecules. This system has attractive features as a peptide nanocarrier for cancer vaccines, including transcutaneous targeting of professional APCs in the skin, high encapsulation efficacy of hydrophilic molecules, and capacity for coloading with a variety of immune stimulants such as adjuvants. We therefore sought to utilize the developed S/O nanodispersion for the delivery of the tyrosine-related protein 2 peptide, TRP-2180-188, as a peptide antigen against melanoma. Transcutaneous vaccination of the S/O nanodispersion coloaded with adjuvant R-848 was associated with a significant inhibition of melanoma growth and suppression of lung metastasis in tumor-bearing mice. Our findings indicate the potential of S/O nanodispersions as an endogenous peptide carrier for cancer vaccines.. |
| 105. |
Muhamad Alif Razi, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya, Formation and characterization of caseinate-chitosan nanocomplexes for encapsulation of Curcumin, JOURNAL OF CHEMICAL ENGINEERING OF JAPAN, 10.1252/jcej.17we293, 51, 5, 445-453, 2018.01, Curcumin holds promise as a therapeutic agent due to its capability of conferring several pharmacological activities. While curcumin shows efficacy in preclinical studies as an anti-cancer agent, its translation into the clinic as a drug has yet to be realized. One possible reason is its poor solubility and stability in water, which decreases the bioavailability. Here, we report the formation of biocompatible nanocomplexes (NCs) from caseinate (CS) and chitosan (CH) using an electrostatic interaction-based approach to stabilize and enhance water solubility of curcumin. The formation of CS-CH NCs (CCNCs) was studied as a function of CH concentration. We show that positively charged NCs, having size between approx. 250 nm with a narrow distribution was formed by adjusting CH concentration. CCNCs successfully entrapped curcumin with a high entrapment efficiency. Curcumin was possibly located in a hydrophobic region of CS as indicated by a blue-shift in the emission maxima of curcumin. Ultimately, the stability and water solubility of curcumin in CCNCs could be remarkably enhanced. These results suggest that CCNCs would be useful for increasing the potential of curcumin as a preventive or therapeutic agent.. |
| 106. |
Akitsu Masuda, Kosuke Minamihata, Masato Hino, Daisuke Morokuma, Noriko Karasaki, Hiroaki Mon, Noriho Kamiya, Takahiro Kusakabe, Man Lee, Modular surface display for porcine circovirus type 2 virus-like particles mediated by microbial transglutaminase, Journal of Insect Biotechnology and Sericology, 10.11416/jibs.87.2_053, 87, 2, 53-60, 2018.01, Virus-like particles (VLPs) are multi-protein complex, which mimic viral particles without viral genomes. Recently various applications of VLPs for medical and pharmaceutical fields are developed. Furthermore, surface modification of VLPs is expected to extend their functional versatility. As an enzymatic strategy for the modification, microbial transglutaminase (MTG) that catalyzes the covalent bond between a glutamine residue and a ly-sine residue or a primary amine is suitable for efficient protein ligation. In the previous study, we demonstrated the efficient production of immunogenic VLPs of porcine circovirus type 2 (PCV2) using silkworm-baculovirus expression vector system (silkworm-BEVS). Herein, we established the display system of exotic molecules to VLPs by enzymatic covalent cross-linking using MTG. For the target modification, Q-tag (YPLQMRG) that is MTG reactive sequence were introduced into the N-terminus, C-terminus, or loop BC of capsid protein of PCV2 and successfully expressed as VLPs in silkworm pupae. Of these, PCV2 VLPs with Q-tag at the loop BC and C-terminus were efficiently conjugated with FITC-cadaverine and K-tagged (MRHKGS) EGFP by MTG. These results showed that flexible surface modification of VLPs mediated by MTG could be used for development of several therapeutic tools such as multivalent vaccines.. |
| 107. |
Lili Jia, Kosuke Minamihata, Hirofumi Ichinose, Kouhei Tsumoto, Noriho Kamiya, Polymeric SpyCatcher Scaffold Enables Bioconjugation in a Ratio-Controllable Manner, Biotechnology Journal, 10.1002/biot.201700195, 12, 12, 2017.12, Conjugating enzymes into a large protein assembly often results in an enhancement of overall catalytic activity, especially when different types of enzymes that work cooperatively are assembled together. However, exploring the proper method to achieve protein assemblies with high stability and also to avoid loss of the function of each component for efficient enzyme clustering is remained challenging. Assembling proteins onto synthetic scaffolds through varied post-translational modification methods is particularly favored since the proteins can be site-specifically conjugated together with less activity loss. Here, a SpyCatcher polymer is prepared through catalytic reaction of horseradish peroxidase (HRP) and serves as a polymeric proteinaceous scaffold for construction of protein assemblies. Taking advantage of the favorable SpyCatcher–SpyTag interaction, SpyTagged proteins can be easily assembled onto the polymeric SpyCatcher scaffold with controllable binding ratio and site specificity. Firstly, the feasibility of construction of ratio-controllable binary artificial hemicellulosomes by assembling endoxylanase and arabinofuranosidase is explored. This construct achieves higher sugar conversion than that of the free enzymes when the proportion of arabinofuranosidase is high, because the close spatial proximity of the enzymes allows them to work in a synergistic manner. Another application for biosensing is developed by conjugating SpyTagged Nanoluc and protein G onto SpyCatcher polymer. Due to the protein clustering effect, an amplified luminescent intensity is achieved by the resulting conjugates than chimera protein of Nanoluc and protein G in ovalbumin detection in ELISA.. |
| 108. |
Mari Takahara, Rie Wakabayashi, Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Primary Amine-Clustered DNA Aptamer for DNA-Protein Conjugation Catalyzed by Microbial Transglutaminase, Bioconjugate Chemistry, 10.1021/acs.bioconjchem.7b00594, 28, 12, 2954-2961, 2017.12, DNA-protein conjugates are promising biomolecules for use in areas ranging from therapeutics to analysis because of the dual functionalities of DNA and protein. Conjugation requires site-specific and efficient covalent bond formation without impairing the activity of both biomolecules. Herein, we have focused on the use of a microbial transglutaminase (MTG) that catalyzes the cross-linking reaction between a glutamine residue and a primary amine. In a model bioconjugation, a highly MTG-reactive Gln (Q)-donor peptide (FYPLQMRG, FQ) was fused to enhanced green fluorescent protein (FQ-EGFP) and a primary amine-clustered DNA aptamer was enzymatically synthesized as a novel acyl-acceptor substrate of MTG, whose combination leads to efficient and convenient preparation of DNA-protein conjugates with high purity. Dual functionality of the obtained DNA-EGFP conjugate was evaluated by discrimination of cancer cells via c-Met receptor recognition ability of the DNA aptamer. The DNA aptamer-EGFP conjugate only showed fluorescence toward cells with c-Met overexpression, indicating the retention of the biochemical properties of the DNA and EGFP in the conjugated form.. |
| 109. |
Lili Jia, Kosuke Minamihata, Hirofumi Ichinose, Kouhei Tsumoto, Noriho Kamiya, Polymeric SpyCatcher Scaffold Enables Bioconjugation in a Ratio-Controllable Manner, BIOTECHNOLOGY JOURNAL, 10.1002/biot.201700195, 12, 12, 2017.12, Conjugating enzymes into a large protein assembly often results in an enhancement of overall catalytic activity, especially when different types of enzymes that work cooperatively are assembled together. However, exploring the proper method to achieve protein assemblies with high stability and also to avoid loss of the function of each component for efficient enzyme clustering is remained challenging. Assembling proteins onto synthetic scaffolds through varied post-translational modification methods is particularly favored since the proteins can be site-specifically conjugated together with less activity loss. Here, a SpyCatcher polymer is prepared through catalytic reaction of horseradish peroxidase (HRP) and serves as a polymeric proteinaceous scaffold for construction of protein assemblies. Taking advantage of the favorable SpyCatcher-SpyTag interaction, SpyTagged proteins can be easily assembled onto the polymeric SpyCatcher scaffold with controllable binding ratio and site specificity. Firstly, the feasibility of construction of ratio-controllable binary artificial hemicellulosomes by assembling endoxylanase and arabinofuranosidase is explored. This construct achieves higher sugar conversion than that of the free enzymes when the proportion of arabinofuranosidase is high, because the close spatial proximity of the enzymes allows them to work in a synergistic manner. Another application for biosensing is developed by conjugating SpyTagged Nanoluc and protein G onto SpyCatcher polymer. Due to the protein clustering effect, an amplified luminescent intensity is achieved by the resulting conjugates than chimera protein of Nanoluc and protein G in ovalbumin detection in ELISA.. |
| 110. |
Mad Takahara, Rie Wakabayashi, Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Primary Amine-Clustered DNA Aptamer for DNA-Protein Conjugation Catalyzed by Microbial Transglutaminase, BIOCONJUGATE CHEMISTRY, 10.1021/acs.bioconjchem.7b00594, 28, 12, 2954-2961, 2017.12, DNA-protein conjugates are promising biomolecules for use in areas ranging from therapeutics to analysis because of the dual functionalities of DNA and protein. Conjugation requires site-specific and efficient covalent bond formation without impairing the activity of both biomolecules. Herein, we have focused on the use of a microbial transglutaminase (MTG) that catalyzes the cross-linking reaction between a glutamine residue and a primary amine. In a model bioconjugation, a highly MTG-reactive Gln (Q)-donor peptide (FYPLQMRG, FQ) was fused to enhanced green fluorescent protein (FQEGFP) and a primary amine clustered DNA aptamer was enzymatically synthesized as a novel aryl-acceptor substrate of MTG, whose combination leads to efficient and convenient preparation of DNA-protein conjugates with high purity. Dual functionality of the obtained DNA-EGFP conjugate was evaluated by discrimination of cancer cells via c-Met receptor recognition ability of the DNA aptamer. The DNA aptamer-EGFP conjugate only showed fluorescence toward cells with c-Met overexpression, indicating the retention of the biochemical properties of the DNA and EGFP in the conjugated form.. |
| 111. |
Qingliang Kong, Momoko Kitaoka, Rie Wakabayashi, Noriho Kamiya, Masahiro Goto, Transcutaneous immunotherapy of pollinosis using solid-in-oil nanodispersions loaded with T cell epitope peptides, International Journal of Pharmaceutics, 10.1016/j.ijpharm.2017.07.020, 529, 1-2, 401-409, 2017.08, Pollinosis, a typical seasonal allergy, is a serious public health problem. Limited numbers of patients receive curative immunotherapy instead of symptomatic therapy; however, there are still some concerns about the inconvenience and side effects of subcutaneous injections and sublingual administration caused by immunotherapy. Here, we propose a simple and safe transcutaneous immunotherapy using solid-in-oil (S/O) nanodispersions loaded with vaccine T cell epitope peptides derived from pollen allergen. S/O nanodispersions are oil-based dispersions of antigens coated with hydrophobic surfactants. They have a high potential to deliver biomolecules including peptides or proteins to immune cells in the skin, and to induce an immune response. The result of quantitative and qualitative analysis by in vitro permeation experiments demonstrated the effective permeation of T cell epitope peptides into the skin. Furthermore, in vivo experiments using a pollinosis mouse model indicated that the S/O nanodispersions loaded with T cell epitopes suppressed serum antibody IgE and cytokine production, and alleviated allergic symptoms to a similar therapeutic level to that observed for subcutaneous injection. These results indicate the potential of transcutaneous immunotherapy using S/O nanodispersions for the future treatment of pollinosis.. |
| 112. |
Qingliang Kong, Momoko Kitaoka, Rie Wakabayashi, Noriho Kamiya, Masahiro Goto, Transcutaneous immunotherapy of pollinosis using solid-in-oil nanodispersions loaded with T cell epitope peptides, INTERNATIONAL JOURNAL OF PHARMACEUTICS, 10.1016/j.ijpharm.2017.07.020, 529, 1-2, 401-409, 2017.08, Pollinosis, a typical seasonal allergy, is a serious public health problem. Limited numbers of patients receive curative immunotherapy instead of symptomatic therapy; however, there are still some concerns about the inconvenience and side effects of subcutaneous injections and sublingual administration caused by immunotherapy. Here, we propose a simple and safe transcutaneous immunotherapy using solid-in-oil (S/O) nanodispersions loaded with vaccine T cell epitope peptides derived from pollen allergen. S/O nanodispersions are oil-based dispersions of antigens coated with hydrophobic surfactants. They have a high potential to deliver biomolecules including peptides or proteins to immune cells in the skin, and to induce an immune response. The result of quantitative and qualitative analysis by in vitro permeation experiments demonstrated the effective permeation of T cell epitope peptides into the skin. Furthermore, in vivo experiments using a pollinosis mouse model indicated that the S/O nanodispersions loaded with T cell epitopes suppressed serum antibody IgE and cytokine production, and alleviated allergic symptoms to a similar therapeutic level to that observed for subcutaneous injection. These results indicate the potential of transcutaneous immunotherapy using S/O nanodispersions for the future treatment of pollinosis. (C) 2017 Elsevier B.V. All rights reserved.. |
| 113. |
Wataru Yoshida, Yuzo Baba, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Extraction and Stripping Behavior of Platinum Group Metals Using an Amic-Acid-Type Extractant, JOURNAL OF CHEMICAL ENGINEERING OF JAPAN, 10.1252/jcej.16we335, 50, 7, 521-526, 2017.07, The recovery and separation of platinum group metals (PGMs) are of considerable significance for metal sustainability. To establish an effcient extraction process for PGMs, the novel extractant N-[N,N-di(2-ethylhexyl)aminocarbonylmethyl] phenylalanine (D2EHAF), containing a phenylalanine moiety as the metal-affinity group, was synthesized. The extraction of PGMs from aqueous HCl and HNO3 solutions with D2EHAF, or our previously developed N-[ N, N-di(2-ethylhexyl) aminocarbonylmethyl]glycine (D2EHAG), in n-dodecane was investigated. Both D2EHAF and D2EHAG exhibited high extraction affinities for Os4+, Pt4+, and Pd2+ in aqueous HCl. In a HNO3 solution, D2EHAF extracted Pd2+ more efficiently than D2EHAG, and the extraction of Pd2+ from a 1 mol dm(-3) HNO3 solution with D2EHAF proceeded quantitatively. The extracted metal ions were stripped from the organic solution using a highly acidic solution, or thiourea.. |
| 114. |
Ryosuke Yamada, Kazunori Nakashima, Nanami Asai-Nakashima, Wataru Tokuhara, Nobuhiro Ishida, Satoshi Katahira, Noriho Kamiya, Chiaki Ogino, Akihiko Kondo, Direct Ethanol Production from Ionic Liquid-Pretreated Lignocellulosic Biomass by Cellulase-Displaying Yeasts, APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY, 10.1007/s12010-016-2322-2, 182, 1, 229-237, 2017.05, Among the many types of lignocellulosic biomass pretreatment methods, the use of ionic liquids (ILs) is regarded as one of the most promising strategies. In this study, the effects of four kinds of ILs for pretreatment of lignocellulosic biomass such as bagasse, eucalyptus, and cedar were evaluated. In direct ethanol fermentation from biomass incorporated with ILs by cellulase-displaying yeast, 1-butyl-3-methylimidazolium acetate ([Bmim][OAc]) was the most effective IL. The ethanol production and yield from [Bmim][OAc]-pretreated bagasse reached 0.81 g/L and 73.4% of the theoretical yield after fermentation for 96 h. The results prove the initial concept, in which the direct fermentation from lignocellulosic biomass effectively promoted by the pretreatment with IL.. |
| 115. |
Tatsukawa, Hideki, Liu, Hong Hong, Oba, Shota, Noriho Kamiya, Nakanishi, Yoichi, Hitomi, Kiyotaka, FRET-based detection of isozyme-specific activities of transglutaminases, AMINO ACIDS, 10.1007/s00726-016-2322-0, 49, 3, 615-623, 2017.03. |
| 116. |
Hideki Tatsukawa, Hong Hong Liu, Shota Oba, Noriho Kamiya, Yoichi Nakanishi, Kiyotaka Hitomi, FRET-based detection of isozyme-specific activities of transglutaminases., Amino acids, 10.1007/s00726-016-2322-0, 49, 3, 615-623, 2017.03, Transglutaminases (TGs) comprise a protein family in which the members catalyze the formation of isopeptide bonds between glutamine and lysine residues in various proteins. Expression studies on its three major members, FXIII, TG1, and TG2, have been performed in a relatively large number of mammalian tissues in comparison with those on the other isozymes. We previously identified a highly reactive substrate peptide, including glutamine, for each isozyme from a phage display library and developed a method for detecting isozyme-specific activities by incorporating a labeled substrate peptide into lysine residues of proteins. Here, we describe genetically encoded Förster resonance energy transfer (FRET)-based probes composed of each fluorescence protein (Cerulean and EVenus) fused with substrate peptides. The probe pairs, designated as Trac-MTG (His-CerΔ11-LQ/EV-K-His) containing linker and substrate peptide sequence for microbial TG (MTG), increased the EVenus:Cerulean fluorescence intensity ratio by more than 1.5-fold. Furthermore, we demonstrated that Trac-TG1 (His-CerΔ11-K5) and Trac-TG2 (His-CerΔ11-T26) containing substrate peptide sequence for mammalian TGs successfully detected the isozyme-specific activity of TG1 and TG2, respectively. In this study, we developed a rapid and convenient experimental system for measuring the isozyme-specific activity of TGs. The application of these probes for analyses in cells and tissues will be helpful for elucidating the physiological and pathological functions of TGs.. |
| 117. |
Kitaoka M, Naritomi A, Kawabe Y, Kamihira M, Kamiya N, Goto M, Transcutaneous pollinosis immunotherapy using a solid-in-oil nanodispersion system carrying T cell epitope peptide and R848., Bioengineering & translational medicine, 10.1002/btm2.10048, 2, 1, 102-108, 2017.03. |
| 118. |
Rie Wakabayashi, Yahiro, Kensuke, Hayashi, Kounosuke, Masahiro Goto, Noriho Kamiya, Protein-Grafted Polymers Prepared Through a Site-Specific Conjugation by Microbial Transglutaminase for an Immunosorbent Assay, BIOMACROMOLECULES, 10.1021/acs.biomac.6b01538, 18, 2, 422-430, 2017.02. |
| 119. |
Rie Wakabayashi, Kensuke Yahiro, Kounosuke Hayashi, Masahiro Goto, Noriho Kamiya, Protein-Grafted Polymers Prepared Through a Site-Specific Conjugation by Microbial Transglutaminase for an Immunosorbent Assay, BIOMACROMOLECULES, 10.1021/acs.biomac.6b01538, 18, 2, 422-430, 2017.02, Protein polymer conjugates have been developed in many fields. Most hybrids are composed of one protein attached to one or several polymer chains. The other form of hybrid involves the construction of multiple proteins on one polymer chain, thereby facilitating protein assemblies that provide multivalent effects. Unfortunately, synthetic methods for production of these types of hybrids are limited and challenging because precise control of the conjugation sites is needed. Herein, a novel synthetic polymer that can enzymatically assemble multiple proteins was developed. Polyacrylamide grafted with multiple microbial transglutaminase (MTG)-recognizable peptide derivatives was synthesized, and MTG-catalyzed site-specific conjugation of proteins with the polymer was achieved. The application for immunological biosensing was demonstrated using the assembly of a fusion protein composed of antibody-binding and enzyme moieties. This enzymatic method to synthesize a one-dimensional protein assembly on a synthetic polymer is versatile and can be expanded to a wide range of applications.. |
| 120. |
Wataru Yoshida, Yuzo Baba, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Extraction and stripping behavior of platinum group metals using an amic-acid-type extractant, JOURNAL OF CHEMICAL ENGINEERING OF JAPAN, 10.1252/jcej.16we335, 50, 7, 521-526, 2017.01, The recovery and separation of platinum group metals (PGMs) are of considerable significance for metal sustainability. To establish an efficient extraction process for PGMs, the novel extractant N-[N,N-di(2-ethylhexyl)aminocarbonylmethyl] phenylalanine (D2EHAF), containing a phenylalanine moiety as the metal-affinity group, was synthesized. The extraction of PGMs from aqueous HCl and HNO3 solutions with D2EHAF, or our previously developed N-[N,N-di(2-ethylhexyl) aminocarbonylmethyl]glycine (D2EHAG), in n-dodecane was investigated. Both D2EHAF and D2EHAG exhibited high extraction affinities for Os4+, Pt4+, and Pd2+ in aqueous HCl. In a HNO3 solution, D2EHAF extracted Pd2+ more efficiently than D2EHAG, and the extraction of Pd2+ from a 1 mol dm-3 HNO3 solution with D2EHAF proceeded quantitatively. The extracted metal ions were stripped from the organic solution using a highly acidic solution, or thiourea.. |
| 121. |
Ryosuke Yamada, Kazunori Nakashima, Nanami Asai-Nakashima, Wataru Tokuhara, Nobuhiro Ishida, Satoshi Katahira, Noriho Kamiya, Chiaki Ogino, Akihiko Kondo, Direct Ethanol Production from Ionic Liquid-Pretreated Lignocellulosic Biomass by Cellulase-Displaying Yeasts, Applied Biochemistry and Biotechnology, 10.1007/s12010-016-2322-2, 182, 1, 229-237, 2017, Among the many types of lignocellulosic biomass pretreatment methods, the use of ionic liquids (ILs) is regarded as one of the most promising strategies. In this study, the effects of four kinds of ILs for pretreatment of lignocellulosic biomass such as bagasse, eucalyptus, and cedar were evaluated. In direct ethanol fermentation from biomass incorporated with ILs by cellulase-displaying yeast, 1-butyl-3-methylimidazolium acetate ([Bmim][OAc]) was the most effective IL. The ethanol production and yield from [Bmim][OAc]-pretreated bagasse reached 0.81 g/L and 73.4% of the theoretical yield after fermentation for 96 h. The results prove the initial concept, in which the direct fermentation from lignocellulosic biomass effectively promoted by the pretreatment with IL.. |
| 122. |
Hideki Tatsukawa, Hong Hong Liu, Shota Oba, Noriho Kamiya, Yoichi Nakanishi, Kiyotaka Hitomi, FRET-based detection of isozyme-specific activities of transglutaminases, Amino Acids, 10.1007/s00726-016-2322-0, 49, 3, 615-623, 2017, Transglutaminases (TGs) comprise a protein family in which the members catalyze the formation of isopeptide bonds between glutamine and lysine residues in various proteins. Expression studies on its three major members, FXIII, TG1, and TG2, have been performed in a relatively large number of mammalian tissues in comparison with those on the other isozymes. We previously identified a highly reactive substrate peptide, including glutamine, for each isozyme from a phage display library and developed a method for detecting isozyme-specific activities by incorporating a labeled substrate peptide into lysine residues of proteins. Here, we describe genetically encoded Förster resonance energy transfer (FRET)-based probes composed of each fluorescence protein (Cerulean and EVenus) fused with substrate peptides. The probe pairs, designated as Trac-MTG (His-CerΔ11-LQ/EV-K-His) containing linker and substrate peptide sequence for microbial TG (MTG), increased the EVenus:Cerulean fluorescence intensity ratio by more than 1.5-fold. Furthermore, we demonstrated that Trac-TG1 (His-CerΔ11-K5) and Trac-TG2 (His-CerΔ11-T26) containing substrate peptide sequence for mammalian TGs successfully detected the isozyme-specific activity of TG1 and TG2, respectively. In this study, we developed a rapid and convenient experimental system for measuring the isozyme-specific activity of TGs. The application of these probes for analyses in cells and tissues will be helpful for elucidating the physiological and pathological functions of TGs.. |
| 123. |
Kitaoka, Momoko, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya, Solid-in-oil nanodispersions for transdermal drug delivery systems, BIOTECHNOLOGY JOURNAL, 10.1002/biot.201600081, 11, 11, 1375-1385, 2016.11. |
| 124. |
Moriyama, Kousuke, Naito, Shono, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya, Enzymatically prepared redox-responsive hydrogels as potent matrices for hepatocellular carcinoma cell spheroid formation, BIOTECHNOLOGY JOURNAL, 10.1002/biot.201600087, 11, 11, 1452-1460, 2016.11. |
| 125. |
Hyung Joon Cha, Noriho Kamiya, Vikineswary Sabaratnam, Editorial
Asian Congress of Biotechnology 2015, Biotechnology Journal, 10.1002/biot.201600650, 11, 11, 1371-1372, 2016.11. |
| 126. |
Kousuke Moriyama, Shono Naito, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya, Enzymatically prepared redox-responsive hydrogels as potent matrices for hepatocellular carcinoma cell spheroid formation, BIOTECHNOLOGY JOURNAL, 10.1002/biot.201600087, 11, 11, 1452-1460, 2016.11, Cellular spheroids have been received much attention in the biological and biomedical fields, especially as a base material for drug assays, regenerative medicine, and tissue engineering. Hydrogels have potential for scalable preparation of spheroids because they provide a spatial environment suitable for three-dimensional cell cultivation. Herein, the potential use of a redox-responsive hydrogel as a scaffold for preparation and recovery of spheroids is reported. A hydrogel composed of poly(ethylene glycol) (PEG), which can be degraded using cysteine as a reducing agent under mild conditions, is prepared by mixing an octa-thiolated PEG derivative (8-arm PEG-SH), horseradish peroxidase and a small phenolic compound (Glycyl-L-tyrosine). Human hepatocellular carcinoma cells (HepG2) are encapsulated in the hydrogel and cellular spheroids formed by proliferation within the scaffolds. After seven days of cultivation, the size of the HepG2 spheroids reached a diameter between approximate to 40 and 60 mu m, depending on the 8-arm PEG-SH concentration. Liver-specific functions of the HepG2 spheroids such as albumin secretion and urea production are retained at higher levels than those of cells prepared from traditional two-dimensional monolayers. These results suggest that the system presented here has potential for preparation of cellular spheroids for tissue engineering applications.. |
| 127. |
Momoko Kitaoka, Rie Wakabayashi, Noriho Kamiya, Masahiro Goto, Solid-in-oil nanodispersions for transdermal drug delivery systems, BIOTECHNOLOGY JOURNAL, 10.1002/biot.201600081, 11, 11, 1375-1385, 2016.11, Transdermal administration of drugs has advantages over conventional oral administration or administration using injection equipment. The route of administration reduces the opportunity for drug evacuation before systemic circulation, and enables long-lasting drug administration at a modest body concentration. In addition, the skin is an attractive route for vaccination, because there are many immune cells in the skin. Recently, solid-in-oil nanodisperison (S/O) technique has demonstrated to deliver cosmetic and pharmaceutical bioactives efficiently through the skin. S/O nanodispersions are nanosized drug carriers designed to overcome the skin barrier. This review discusses the rationale for preparation of efficient and stable S/O nanodispersions, as well as application examples in cosmetic and pharmaceutical materials including vaccines. Drug administration using a patch is user-friendly, and may improve patient compliance. The technique is a potent transcutaneous immunization method without needles.. |
| 128. |
Jia, Lili, Budinova, Geisa A. L. G., Takasugi, Yusaku, Noda, Shuhei, Tanaka, Tsutomu, Hirofumi Ichinose, Masahiro Goto, Kamiya, Noriho, Synergistic degradation of arabinoxylan by free and immobilized xylanases and arabinofuranosidase, BIOCHEMICAL ENGINEERING JOURNAL, 10.1016/j.bej.2016.07.013, 114, 271-278, 2016.10. |
| 129. |
Takahara, Mari, Budinova, Geisa Aparecida Lopes Goncalves, Nakazawa, Hikaru, Mori, Yutaro, Umetsu, Mitsuo, Kamiya, Noriho, Salt-Switchable Artificial Cellulase Regulated by a DNA Aptamer, BIOMACROMOLECULES, 10.1021/acs.biomac.6b01141, 17, 10, 3356-3362, 2016.10. |
| 130. |
Mari Takahara, Geisa Aparecida Lopes Goncalves Budinova, Hikaru Nakazawa, Yutaro Mori, Mitsuo Umetsu, Noriho Kamiya, Salt-Switchable Artificial Cellulase Regulated by a DNA Aptamer, BIOMACROMOLECULES, 10.1021/acs.biomac.6b01141, 17, 10, 3356-3362, 2016.10, A novel artificial cellulase was developed by conjugating a DNA aptamer to an endoglucanase catalytic domain, thereby substituting the natural carbohydrate-binding module. Circular dichroism spectroscopy and adsorption isotherm showed the binding motif of cellulose-binding DNA aptamer (CelApt) was G-quadruplex and stem-loop structures stabilized in the presence of salts, and CelApt binding preferred the amorphous region of the solid cellulose. By introducing the revealed salt-switchable cellulose binding nature of CelApt into a catalytic domain of a cellulase, we created CelApt-catalytic domain conjugate possessing both controllable adsorption on the solid substrates and equal enzymatic activity to the wild-type cellulase. Thus potential use of a responsive DNA aptamer for biocatalysis at a solid surface was demonstrated.. |
| 131. |
Lili Jia, Geisa A. L. G. Budinova, Yusaku Takasugi, Shuhei Noda, Tsutomu Tanaka, Hirofumi Ichinose, Masahiro Goto, Noriho Kamiya, Synergistic degradation of arabinoxylan by free and immobilized xylanases and arabinofuranosidase, BIOCHEMICAL ENGINEERING JOURNAL, 10.1016/j.bej.2016.07.013, 114, 271-278, 2016.10, Effective degradation of hemicellulose is of utmost importance in a wide variety of applications in bioindustry. Five endoxylanases from different glycoside hydrolase families and microorganisms were tested with an arabinofuranosidase, Araf51A, for the hydrolysis of insoluble wheat arabinoxylan, which is a structural component of hemicellulose. The optimized combination was XynZ/Xyn11A/Araf51A with a loading ratio of 2:2:1, and the value of degree of synergy increased with the increase of Araf51A proportion in the enzyme mixture. Afterwards, selected enzymes were immobilized on commercial magnetic nanoparticles through covalent bonding. Both free and immobilized enzymes showed a similar conversion to reducing sugars after hydrolysis for 48 h. After 10 cycles, approximately 20% of the initial enzymatic activity of both the individual or mixture of immobilized enzymes was retained. A 5.5-fold increase in the production of sugars was obtained with a mixture of enzymes immobilized after 10 cycles in total compared with free enzymes. Importantly, a sustainable synergism between immobilized arabinofuranosidase and immobilized endoxylanases in the hydrolysis of arabinoxylan was demonstrated. (C) 2016 Elsevier B.V. All rights reserved.. |
| 132. |
Hosomomi, Yukiho, F. Kubota, Rie Wakabayashi, Kamiya, Noriho, Masahiro Goto, Diglycolic amic acid-modified E. coli as a biosorbent for the recovery of rare earth elements, BIOCHEMICAL ENGINEERING JOURNAL, 10.1016/j.bej.2016.06.005, 113, 102-106, 2016.09. |
| 133. |
Furukawa, Shin-ya, Hattori, Gaku, Sakai, Shinji, Kamiya, Noriho, Highly efficient and low toxic skin penetrants composed of amino acid ionic liquids, RSC Adv., 10.1039/C6RA16926K, 6, 87753-87755, 2016.09. |
| 134. |
Diglycolic amic acid-modified E. coli as a biosorbent for the recovery of rare earth elements
© 2016 Elsevier B.V. Biosorption has recently attracted much attention as an alternative to conventional techniques for the recovery of rare earth elements (REEs). In this study, Escherichia coli (E. coli) was chemically modified to improve its performance as a biosorbent for REEs. The diglycolic amic acid group, which shows high affinity to REEs, was introduced by succinylation of the amine groups on the E. coli. Adsorption curves using the modified E. coli were characteristic of the diglycolic amic acid group. The adsorption performance for transition metal ions was not affected by the modification. These results suggest that modification of E. coli with a functional group with high affinity to REEs increases the effectiveness of adsorption. The maximum uptakes of REEs on the modified E. coli were doubled. Modification of E. coli is an effective method for enhancing the adsorption performance for REEs.. |
| 135. |
Uju, Masahiro Goto, Noriho Kamiya, Powerful peracetic acid-ionic liquid pretreatment process for the efficient chemical hydrolysis of lignocellulosic biomass, BIORESOURCE TECHNOLOGY, 10.1016/j.biortech.2016.04.121, 214, 487-495, 2016.08. |
| 136. |
Kazunori Matsuura, Tomohiro Nakamura, Kenta Watanabe, Takanori Noguchi, Kosuke Minamihata, Kamiya, Noriho, Nobuo Kimizuka, Self-assembly of Ni-NTA-modified beta-annulus peptides into artificial viral capsids and encapsulation of His-tagged proteins, Org. Biomol. Chem., 10.1039/C6OB01227B, 14, 7869-7874, 2016.08. |
| 137. |
Powerful peracetic acid-ionic liquid pretreatment process for the efficient chemical hydrolysis of lignocellulosic biomass
© 2016 Elsevier Ltd. The aim of this work was to design a new method for the efficient saccharification of lignocellulosic biomass (LB) using a combination of peracetic acid (PAA) pretreatment with ionic liquid (IL)-HCl hydrolysis. The pretreatment of LBs with PAA disrupted the lignin fractions, enhanced the dissolution of LB and led to a significant increase in the initial rate of the IL-HCl hydrolysis. The pretreatment of Bagasse with PAA prior to its 1-buthyl-3-methylimidazolium chloride ([Bmim][Cl])-HCl hydrolysis, led to an improvement in the cellulose conversion from 20% to 70% in 1.5 h. Interestingly, the 1-buthyl-3-methylpyridium chloride ([Bmpy][Cl])-HCl hydrolysis of Bagasse gave a cellulose conversion greater than 80%, with or without the PAA pretreatment. For LB derived from seaweed waste, the cellulose conversion reached 98% in 1 h. The strong hydrolysis power of [Bmpy][Cl] was attributed to its ability to transform cellulose I to II, and lowering the degree of polymerization of cellulose.. |
| 138. |
Masato Hino, Takuji Kawanami, Noriho Kamiya, Tsuneyuki Tatsuke, Yutaka Banno, Takahiro Kusakabe, JAE MAN LEE, High-level expression and purification of biologically active human IL-2 using silkworm-baculovirus expression vector system, JOURNAL OF ASIA-PACIFIC ENTOMOLOGY, 10.1016/j.aspen.2016.03.014, 19, 2, 313-317, 2016.06. |
| 139. |
Takahara, Mari, Hayashi, Kounosuke, Masahiro Goto, Kamiya, Noriho, Enzymatic conjugation of multiple proteins on a DNA aptamer in a tail-specific manner, BIOTECHNOLOGY JOURNAL, 10.1002/biot.201500560, 11, 6, 814-823, 2016.06. |
| 140. |
Enzymatic conjugation of multiple proteins on a DNA aptamer in a tail-specific manner
Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim Conjugation of single-strand DNA aptamers and enzymes has been of great significance in bioanalytical and biomedical applications because of the unlimited functions provided by DNA aptamer direction. Therefore, we developed efficient tailing of a DNA aptamer, with end-specific conjugation of multiple enzymes, through enzymatic catalysis. Terminal deoxynucleotidyl transferase (TdT) added multiple Z-Gln-Gly (Z-QG) moieties to the 3′-end of a DNA aptamer via the addition of Z-QG-modified deoxyuridine triphosphate (Z-QG-dUTP) and deoxynucleoside triphosphates (dNTPs). The resultant (Z-QG)m-(dN)l-aptamer, whose Z-QGs with dN spacers served as stickers for microbial transglutaminase (MTG), were crosslinked between the Z-QGs on the DNA and a substrate peptide sequence containing lysine (K), fused to a recombinant enzyme (i.e. bacterial alkaline phosphatase; BAP) by MTG. The incorporation efficiency of Z-QG moieties on the aptamer tail and the subsequent conjugation efficiency with multiple enzyme molecules were dramatically altered by the presence of dNTPs, revealing that a combination of Z-QG-dUTP/dTTP comprised the best labeling efficiency and corresponding properties during analytical performance. Thus, a novel optimized platform for designing (BAP)n-(dT)l-DNA aptamers was demonstrated for the first time in this article, offering unique opportunities for tailoring new types of covalent protein-nucleic acid conjugates in a controllable way.. |
| 141. |
High-level expression and purification of biologically active human IL-2 using silkworm-baculovirus expression vector system. |
| 142. |
Akira Tsuchiya, Siti Norulhuda Hashim, Ise, Shoko, Furuhata, Takafumi, Rie Wakabayashi, Masahiro Goto, Kawai, Kiyohiko, Noriho Kamiya, Shinsuke Sando, BODIPY-labeled Fluorescent Aptamer Sensors for Turn-on Sensing of Interferon-gamma and Adenine Compounds on Cells, ANALYTICAL SCIENCES, 32, 5, 543-547, 2016.05. |
| 143. |
Mori, Yutaro, Budinova, Geisa Aparecida Lopes Goncalves, Nakazawa, Hikaru, Umetsu, Mitsuo, Kamiya, Noriho, One-dimensional assembly of functional proteins: toward the design of an artificial cellulosome, Mol. Syst. Des. Eng., 10.1039/C6ME90005D, 1, 1, 66-73, 2016.04, In biological systems, proteins can form well-organized, higher-order structures with unique functions that would be difficult to achieve with a single protein. These proteinaceous supramolecular structures form by self-assembly, and the spatial arrangement of the protein building blocks in them is very important. In the present study, an artificial system was developed using recombinant proteins as building blocks, which were assembled in a one-dimensional manner. The assembly of these building blocks was based on the avidin-biotin interaction. A tetrameric biotin ligand unit was designed so that the 1:4 stoichiometry of the avidin-biotin interaction was altered to a 1:2 directional interaction between the streptavidin and tetrabiotinylated protein units. In a proof-of-concept study, site-specifically tetrabiotin-labeled endoglucanase and cellulose-binding module units were prepared, then these components were self-assembled by mixing with streptavidin to mimic a natural cellulosome. The formation of one-dimensional assemblies of the protein units depended on the stoichiometry of the avidin-biotin interaction sites in the system. Interestingly, the saccharification efficiency improved when the component ratio of protein units in the assemblies was changed.. |
| 144. |
Yukiho HOSOMOMI, Teppei NIIDE, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya, Biocatalytic Formation of Gold Nanoparticles Decorated with Functional Proteins inside Recombinant Escherichia coli Cells, ANALYTICAL SCIENCES, 32, 3, 295-300, Cover page illustration, 2016.03. |
| 145. |
Shin Ya Furukawa, Gaku Hattori, Shinji Sakai, Noriho Kamiya, Highly efficient and low toxic skin penetrants composed of amino acid ionic liquids, RSC Advances, 10.1039/c6ra16926k, 6, 90, 87753-87755, 2016.01, Herein, we introduce an amino acid ester (AAE) as a potent biocompatible counter ion to formulate ionic liquefied active pharmaceutical ingredients. By equimolar pairing of a non-steroidal anti-inflammatory drug (NSAID) as the anion and the AAE as the cation, the liquid formulation was feasibly obtained. In vitro drug permeation results using pig skin revealed a significant enhancement of the transdermal delivery of the ionic liquefied NSAIDs. The AAE showed much lower cytotoxicity to mouse fibroblast L929 cells when compared with that of conventional counter cations, indicating the significant potential of the AAE in the development of new ionic liquid drug formulations.. |
| 146. |
Kazunori Matsuura, Tomohiro Nakamura, Kenta Watanabe, Takanori Noguchi, Kosuke Minamihata, Noriho Kamiya, Nobuo Kimizuka, Self-assembly of Ni-NTA-modified β-annulus peptides into artificial viral capsids and encapsulation of His-tagged proteins, Organic and Biomolecular Chemistry, 10.1039/c6ob01227b, 14, 33, 7869-7874, 2016.01, β-Annulus peptides bearing Cys at the N-terminal from tomato bushy stunt virus were synthesised using a standard Fmoc-protected solid-phase method, and the peptide was modified with Ni-NTA at the N-terminal. The Ni-NTA-modified β-annulus peptide self-assembled into virus-like nanocapsules of approximately 40 nm in diameter. The critical aggregation concentration of these nanocapsules in 10 mM Tris-HCl buffer (pH 7.3) at 25 °C was 0.053 μM, which is 470 times lower than that of unmodified β-annulus peptides. Moreover, size exclusion chromatography of the peptide assembly indicated encapsulation of His-tagged green fluorescent protein in the Ni-NTA-modified artificial viral capsid.. |
| 147. |
Noriho Kamiya, Yutaro Mori, Substrate engineering of microbial transglutaminase for site-specific protein modification and bioconjugation, Transglutaminases
Multiple Functional Modifiers and Targets for New Drug Discovery, 10.1007/978-4-431-55825-5_17, 373-383, 2016.01, Microbial transglutaminase (MTG), a robust enzyme developed initially for the manipulation of edible proteins in the food industry, has now been widely recognized as a practical protein-modifying reagent in the range of biotechnological applications. In this chapter, we introduce the potential use of MTG through our basic studies on the design of novel glutamine (Gln) donor substrates for lysine (Lys)-specific protein modification. Based on the core structure of a conventional transglutaminase substrate, benzyloxycarbonyl-L-glutaminylglycine (Z-QG), new Gln-donor substrates have been developed for the conjugation of recombinant proteins with different functionalities. The first target site for the substrate engineering was the C-terminal carboxylic group of Z-QG, which is feasibly labeled with functional moieties. For the preparation of protein-nucleic acid conjugates with novel molecular architecture, a new nucleotidyl substrate, Z-QG-(d)UTP, was created. We have also explored substitution of the N-terminal protecting group (Z) with fluorophores and biotin, and found that MTG accepts diverse functional groups at the N-terminus by inserting a short linker, leading to an increase in the utility of MTG in site-specific modification of functional proteins. Our results demonstrated how the design of (small) Gln-donor substrates of MTG can expand the scope of enzymatic manipulation in biomolecular engineering.. |
| 148. |
BODIPY-labeled fluorescent aptamer sensors for turn-on sensing of interferon-gamma and adenine compounds on cells
© The Japan Society for Analytical Chemistry. An on-cell aptamer sensor has the potential to reveal cell-cell communications by signalling molecules. We attempted to design new fluorescent aptamer sensors for the sensing of IFN-? and adenine compounds on cells. BODIPY-labeled external quencher-free aptamer sensors have allowed a turn-on detection of the target molecule with improved off/on efficiency.. |
| 149. |
Biocatalytic formation of gold nanoparticles decorated with functional proteins inside recombinant Escherichia coli cells
© The Japan Society for Analytical Chemistry. A novel strategy for the preparation of protein-decorated gold nanoparticles (Au NPs) was developed inside Escherichia coli cells, where an artificial oxidoreductase, composed of antibody-binding protein (pG), Bacillus stearothermophilus glycerol dehydrogenase (BsGLD) and a peptide tag with gold-binding affinity (H6C), was overexpressed in the cytoplasm. In situ formation of Au NPs was promoted by a natural electron-donating cofactor, nicotinamide adenine dinucleotide (NAD), which was regenerated to the reduced form of NADH by the catalytic activity of the fusion protein (pG-BsGLDH6C) overexpressed in the cytoplasm of E. coli, with the concomitant addition of exogenous glycerol to the reaction system. The fusion protein was self-immobilized on Au NPs inside the E. coli cells, which was confirmed by SDS-PAGE and western blotting analyses of the resultant Au NPs. Finally, the IgG binding ability of the pG moiety displayed on Au NPs was evaluated by an enzyme-linked immunosorbent assay. 2016. |
| 150. |
Mutual separation of indium, gallium, and zinc with the amic acid-type extractant D2EHAG containing glycine and amide moieties
Solvent extraction of indium (In3+), gallium (Ga3+), and zinc (Zn2+) was investigated using N-[N,N-di(2-ethylhexyl)aminocarbonylmethyl]glycine (D2EHAG) and N-[N,N-di(2-ethylhexyl)aminocarbonylmethyl]-sarcosine (D2EHAS) which we have developed recently. Indium and gallium were selectively extracted from zinc under high acid conditions (0 > pH ≥ 2.0), and easily stripped using an acidic solution such as 2 mol dm-3 HNO3. The extraction behavior was compared to that of N,N-dioctyldiglycol amic acid (DODGAA), which has a similar molecular structure to D2EHAG, a commercial alkyl monocarboxylic acid extractant, neodecanoic acid (Versatic 10), or an organophosphorus extractant, di(2-ethylhexyl) phosphoric acid (D2EHPA). Based on the results, D2EHAG and D2EHAS were found to be useful for the separation of In3+ and Ga3+ from Zn2+. The extraction mechanisms of these ions with D2EHAG were investigated through slope analysis and loading tests.. |
| 151. |
Selective extraction of scandium from transition metals by synergistic extraction with 2-thenoyltrifluoroacetone and tri-n-octylphosphine oxide
There has been an increasing demand for scandium (Sc). However the supply is insufficient because the separation and recovery of Sc as a byproduct from ores is difficult. In this study, extraction and separation of Sc3+from a sulfuric acid solution containing transition metals such as Al3+, Fe3+, Mn2+, Co2+, Ni2+and Zn2+was investigated using a β-diketone, HTTA and a neutral extractant, TOPO. Due to the synergistic effect with the two extractants, Sc3+was effectively and selectively separated even from Fe3+, the separation of which is difficult with HTTA alone. The extraction mechanism was examined and it was revealed that Sc3+is extracted with three HTTA and one TOPO molecules. Scandium was readily stripped from the extracting phase by 1 M sulfuric acid. The potential use of the combination of the β-diketone with TOPO for the recovery of Sc3+in a refining process was demonstrated.. |
| 152. |
Self-assembly of Ni-NTA-modified β-annulus peptides into artificial viral capsids and encapsulation of His-tagged proteins
© The Royal Society of Chemistry 2016. β-Annulus peptides bearing Cys at the N-terminal from tomato bushy stunt virus were synthesised using a standard Fmoc-protected solid-phase method, and the peptide was modified with Ni-NTA at the N-terminal. The Ni-NTA-modified β-annulus peptide self-assembled into virus-like nanocapsules of approximately 40 nm in diameter. The critical aggregation concentration of these nanocapsules in 10 mM Tris-HCl buffer (pH 7.3) at 25 °C was 0.053 μM, which is 470 times lower than that of unmodified β-annulus peptides. Moreover, size exclusion chromatography of the peptide assembly indicated encapsulation of His-tagged green fluorescent protein in the Ni-NTA-modified artificial viral capsid.. |
| 153. |
Noriho Kamiya, Yutaro Mori, Substrate engineering of microbial transglutaminase for site-specific protein modification and bioconjugation, Transglutaminases: Multiple Functional Modifiers and Targets for New Drug Discovery, 10.1007/978-4-431-55825-5_17, 373-383, 2016.01, Microbial transglutaminase (MTG), a robust enzyme developed initially for the manipulation of edible proteins in the food industry, has now been widely recognized as a practical protein-modifying reagent in the range of biotechnological applications. In this chapter, we introduce the potential use of MTG through our basic studies on the design of novel glutamine (Gln) donor substrates for lysine (Lys)-specific protein modification. Based on the core structure of a conventional transglutaminase substrate, benzyloxycarbonyl-L-glutaminylglycine (Z-QG), new Gln-donor substrates have been developed for the conjugation of recombinant proteins with different functionalities. The first target site for the substrate engineering was the C-terminal carboxylic group of Z-QG, which is feasibly labeled with functional moieties. For the preparation of protein-nucleic acid conjugates with novel molecular architecture, a new nucleotidyl substrate, Z-QG-(d)UTP, was created. We have also explored substitution of the N-terminal protecting group (Z) with fluorophores and biotin, and found that MTG accepts diverse functional groups at the N-terminus by inserting a short linker, leading to an increase in the utility of MTG in site-specific modification of functional proteins. Our results demonstrated how the design of (small) Gln-donor substrates of MTG can expand the scope of enzymatic manipulation in biomolecular engineering.. |
| 154. |
Akira Tsuchiya, Siti N. Hashim, Shoko Ise, Takafumi Furuhata, Kiyohiko Kawai, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya, Shinsuke Sando, BODIPY-labeled fluorescent aptamer sensors for turn-on sensing of interferon-gamma and adenine compounds on cells, analytical sciences, 10.2116/analsci.32.543, 32, 5, 543-547, 2016, An on-cell aptamer sensor has the potential to reveal cell-cell communications by signalling molecules. We attempted to design new fluorescent aptamer sensors for the sensing of IFN-? and adenine compounds on cells. BODIPY-labeled external quencher-free aptamer sensors have allowed a turn-on detection of the target molecule with improved off/on efficiency.. |
| 155. |
Yukiho Hosomomi, Teppei Niide, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya, Biocatalytic formation of gold nanoparticles decorated with functional proteins inside recombinant Escherichia coli cells, analytical sciences, 10.2116/analsci.32.295, 32, 3, 295-300, 2016, A novel strategy for the preparation of protein-decorated gold nanoparticles (Au NPs) was developed inside Escherichia coli cells, where an artificial oxidoreductase, composed of antibody-binding protein (pG), Bacillus stearothermophilus glycerol dehydrogenase (BsGLD) and a peptide tag with gold-binding affinity (H 6 C), was overexpressed in the cytoplasm. In situ formation of Au NPs was promoted by a natural electron-donating cofactor, nicotinamide adenine dinucleotide (NAD), which was regenerated to the reduced form of NADH by the catalytic activity of the fusion protein (pG-BsGLDH 6 C) overexpressed in the cytoplasm of E. coli, with the concomitant addition of exogenous glycerol to the reaction system. The fusion protein was self-immobilized on Au NPs inside the E. coli cells, which was confirmed by SDS-PAGE and western blotting analyses of the resultant Au NPs. Finally, the IgG binding ability of the pG moiety displayed on Au NPs was evaluated by an enzyme-linked immunosorbent assay. 2016. |
| 156. |
Yuzo Baba, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Mutual separation of indium, gallium, and zinc with the amic acid-type extractant D2EHAG containing glycine and amide moieties, Solvent Extraction Research and Development, 10.15261/serdj.23.9, 23, 1, 9-18, 2016, Solvent extraction of indium (In3+), gallium (Ga3+), and zinc (Zn2+) was investigated using N-[N,N-di(2-ethylhexyl)aminocarbonylmethyl]glycine (D2EHAG) and N-[N,N-di(2-ethylhexyl)aminocarbonylmethyl]-sarcosine (D2EHAS) which we have developed recently. Indium and gallium were selectively extracted from zinc under high acid conditions (0 > pH ≥ 2.0), and easily stripped using an acidic solution such as 2 mol dm-3 HNO3. The extraction behavior was compared to that of N,N-dioctyldiglycol amic acid (DODGAA), which has a similar molecular structure to D2EHAG, a commercial alkyl monocarboxylic acid extractant, neodecanoic acid (Versatic 10), or an organophosphorus extractant, di(2-ethylhexyl) phosphoric acid (D2EHPA). Based on the results, D2EHAG and D2EHAS were found to be useful for the separation of In3+ and Ga3+ from Zn2+. The extraction mechanisms of these ions with D2EHAG were investigated through slope analysis and loading tests.. |
| 157. |
Y. Mori, H. Nakazawa, G. A.L. Gonçalves, T. Tanaka, M. Umetsu, N. Kamiya, One-dimensional assembly of functional proteins
Toward the design of an artificial cellulosome, Molecular Systems Design and Engineering, 10.1039/c5me00011d, 1, 1, 66-73, 2016, In biological systems, proteins can form well-organized, higher-order structures with unique functions that would be difficult to achieve with a single protein. These proteinaceous supramolecular structures form by self-assembly, and the spatial arrangement of the protein building blocks in them is very important. In the present study, an artificial system was developed using recombinant proteins as building blocks, which were assembled in a one-dimensional manner. The assembly of these building blocks was based on the avidin-biotin interaction. A tetrameric biotin ligand unit was designed so that the 1:4 stoichiometry of the avidin-biotin interaction was altered to a 1:2 directional interaction between the streptavidin and tetrabiotinylated protein units. In a proof-of-concept study, site-specifically tetrabiotin-labeled endoglucanase and cellulose-binding module units were prepared, and then these components were self-assembled by mixing with streptavidin to mimic a natural cellulosome. The formation of one-dimensional assemblies of the protein units depended on the stoichiometry of the avidin-biotin interaction sites in the system. Interestingly, the saccharification efficiency improved when the component ratio of protein units in the assemblies was changed. The presence of the optimal ratio of the building blocks implies the modularity of the present protein assembly system.. |
| 158. |
Zhigang Zhao, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Selective extraction of scandium from transition metals by synergistic extraction with 2-thenoyltrifluoroacetone and tri-n-octylphosphine oxide, Solvent Extraction Research and Development, 10.15261/serdj.23.137, 23, 2, 137-143, 2016, There has been an increasing demand for scandium (Sc). However the supply is insufficient because the separation and recovery of Sc as a byproduct from ores is difficult. In this study, extraction and separation of Sc3+ from a sulfuric acid solution containing transition metals such as Al3+, Fe3+, Mn2+, Co2+, Ni2+ and Zn2+ was investigated using a β-diketone, HTTA and a neutral extractant, TOPO. Due to the synergistic effect with the two extractants, Sc3+ was effectively and selectively separated even from Fe3+, the separation of which is difficult with HTTA alone. The extraction mechanism was examined and it was revealed that Sc3+ is extracted with three HTTA and one TOPO molecules. Scandium was readily stripped from the extracting phase by 1 M sulfuric acid. The potential use of the combination of the β-diketone with TOPO for the recovery of Sc3+ in a refining process was demonstrated.. |
| 159. |
Siti Norulhuda Hashim, Akira Tsuchiya, Noriho Kamiya, Shinsuke Sando, A Single Fluorophore-labeled Aptamer Sensor for the Detection of Interferon Gamma, Chem. Lett., 44, 12, 1670-1672, 2015.12. |
| 160. |
Siti Norulhuda Hashim, Akira Tsuchiya, Noriho Kamiya, Shinsuke Sando, A Single Fluorophore-labeled Aptamer Sensor for the Detection of Interferon Gamma, CHEMISTRY LETTERS, 10.1246/cl.150794, 44, 12, 1670-1672, 2015.12, Interferon gamma (IFN-gamma) is a cytokine that is mainly secreted from immune cells. It plays an important role in regulating neighboring cellular activities through interactions with cell membrane proteins. Because of its biological importance, there has been increasing demand for a biosensor that can respond to IFN-gamma. Here, we report a fluorescent aptamer sensor that targets IFN-gamma. The 25-nt IFN-gamma aptamer was converted into a turn-on fluorescent sensor by labeling the 3'-end with fluorescein. The designed sensor achieved fluorescence sensing of IFN-gamma with nanomolar affinity.. |
| 161. |
Momoko Kitaoka, Yoko Shin, Noriho Kamiya, Noriho Kamiya, Yoshinori Kawabe, Masamichi Kamihira, Masahiro Goto, Masahiro Goto, Transcutaneous Peptide Immunotherapy of Japanese Cedar Pollinosis Using Solid-in-Oil Nanodispersion Technology, AAPS PharmSciTech, 10.1208/s12249-015-0333-x, 16, 6, 1418-1424, 2015.12, © 2015, American Association of Pharmaceutical Scientists. Peptide immunotherapy is an attractive approach to relieve allergic symptoms such as rhinitis and asthma. Treatment of Japanse cedar pollinosis (Cryptomeria japonica; Cj), from which over one quarter of Japanese population suffer, is becoming a great concern. Recently, oral feeding of a peptide (7crp) consisting of seven immunodominant human T cell epitopes derived from two enzymes present in Cj pollen was demonstrated to have a benefit in treating Cj pollinosis. In this work, we aimed to apply a novel transcutaneous administration system as a simple and easy peptide delivery for an immunotherapy using a T cell epitope peptide. A modified 7crp peptide (7crpR) which contained triarginine linkers between each epitopes was designed to increase water solubility and was encapsulated in a unique solid-in-oil (S/O) nanodispersion. The S/O nanodispersion consists of a nano-sized peptide-surfactant complex dispersed in an oil vehicle. The S/O nanopartilces having an average diameter of 230 nm facilitated the permeation of the peptide 7crpR into the skin and suppressed serum total IgE and antigen-specific IgE levels in a Cj pollinosis mouse model. Transcutaneous administration of the T cell epitope peptide using the S/O nanodispersion system has potential for future simple and easy immunotherapy of Cj pollinosis.. |
| 162. |
Hayashi, Kounosuke, JAE MAN LEE, Tomozoe, Yusuke, takahiro kusakabe, 神谷 典穂, Heme precursor injection is effective for Arthromyces ramosus peroxidase fusion protein production by a silkworm expression system, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 10.1016/j.jbiosc.2015.02.013, 120, 4, 384-386, 2015.10. |
| 163. |
Uju, Wijayanta, Agung Tri, Masahiro Goto, 神谷 典穂, Great potency of seaweed waste biomass from the carrageenan industry for bioethanol production by peracetic acid-ionic liquid pretreatment, BIOMASS & BIOENERGY, 10.1016/j.biombioe.2015.05.023, 81, 63-69, 2015.10. |
| 164. |
Great potency of seaweed waste biomass from the carrageenan industry for bioethanol production by peracetic acid-ionic liquid pretreatment
© 2015 Published by Elsevier Ltd. Seaweed waste biomass from the carrageenan industry (SWBC) is a potential biomass feedstock for producing sustainable biofuel because it increases the product value and reduces the pollutant risk. Peracetic acid (PAA) followed by ionic liquid (IL) pretreatment has been used to increase the enzymatic saccharification of pretreated SWBC. The SWBC cellulose content was comparable with that of terrestrial biomasses. PAA+1-hexylpyridinium chloride ([Hpy][Cl]), and PAA+1-ethyl-3-methylimidazolium diethylphosphate ([Emim][DEP]) pretreatments produced saccharide and unknown oligosaccharide fractions in regenerated water (~4-6% SWBC cellulose content). For 48h of saccharification, the untreated SWBC and the SWBC pretreated using PAA followed by [Hpy][Cl], [Emim][DEP] or 1-ethyl-3-methylimidazole acetate ([Emim][OAc]) produced cellulose conversions of 77, 91, 84 and 62%, respectively. The untreated SWBC had a high cellulose conversion, which may be caused by the low lignin and hemicellulose contents of the SWBC. PAA+IL pretreatment could yield pretreated SWBCs with more amorphous cellulose structures, which lead to an almost-complete cellulose conversion.. |
| 165. |
Heme precursor injection is effective for Arthromyces ramosus peroxidase fusion protein production by a silkworm expression system. |
| 166. |
Yuya Hirakawa, Rie Wakabayashi, Ayaka Naritomi, Masato Sakuragi, Noriho Kamiya, Masahiro Goto, Transcutaneous immunization against cancer using solid-in-oil nanodispersions, MedChemComm, 10.1039/c5md00168d, 6, 7, 1387-1392, 2015.07, Transcutaneous immunization is a novel, non-invasive alternative to conventional immunization by injection. Skin immunocompetence composed of abundant antigen-presenting cells in the epidermal and dermal layers of the skin can provide an effective tool for transcutaneous immunization, whereas the outermost, hydrophobic layer of skin, the stratum corneum, hinders the penetration of antigens into the skin. To realize effective transcutaneous delivery of antigens, we have developed a solid-in-oil (S/O) technique that produces an oil dispersion of hydrophilic biomolecules. In this study, we applied the S/O nanodispersions in transcutaneous immunization to induce cancer immunity. The topical application of S/O nanodispersions bearing ovalbumin (OVA) as a model cancer-antigen allowed the penetration of OVA into the deep dermis region of the skin by intercellular and transfollicular pathways. Inhibition of OVA-bearing tumor growth and production of cytokines responsible for cellular immunity were achieved, demonstrating the applicability of S/O nanodispersions to the induction of cancer immunity.. |
| 167. |
Transcutaneous immunization against cancer using solid-in-oil nanodispersions
© The Royal Society of Chemistry. Transcutaneous immunization is a novel, non-invasive alternative to conventional immunization by injection. Skin immunocompetence composed of abundant antigen-presenting cells in the epidermal and dermal layers of the skin can provide an effective tool for transcutaneous immunization, whereas the outermost, hydrophobic layer of skin, the stratum corneum, hinders the penetration of antigens into the skin. To realize effective transcutaneous delivery of antigens, we have developed a solid-in-oil (S/O) technique that produces an oil dispersion of hydrophilic biomolecules. In this study, we applied the S/O nanodispersions in transcutaneous immunization to induce cancer immunity. The topical application of S/O nanodispersions bearing ovalbumin (OVA) as a model cancer-antigen allowed the penetration of OVA into the deep dermis region of the skin by intercellular and transfollicular pathways. Inhibition of OVA-bearing tumor growth and production of cytokines responsible for cellular immunity were achieved, demonstrating the applicability of S/O nanodispersions to the induction of cancer immunity.. |
| 168. |
Lili Jia, LOPES GONCALVES GEISA APARECIDA, Yusaku Takasugi, Yutaro Mori, Shuhei Noda, Tsutomu Tanaka, Hirofumi Ichinose, 神谷 典穂, Effect of pretreatment methods on the synergism of cellulase and xylanase during the hydrolysis of bagasse, BIORESOURCE TECHNOLOGY, 10.1016/j.biortech.2015.02.041, 185, 158-164, 2015.06. |
| 169. |
Lili Jia, Geisa A.L. Goncalves, Yusaku Takasugi, Yutaro Mori, Shuhei Noda, Tsutomu Tanaka, Hirofumi Ichinose, Noriho Kamiya, Effect of pretreatment methods on the synergism of cellulase and xylanase during the hydrolysis of bagasse, Bioresource Technology, 10.1016/j.biortech.2015.02.041, 185, 158-164, 2015.06. |
| 170. |
LOPES GONCALVES GEISA APARECIDA, Yusaku Takasugi, Lili Jia, Yutaro Mori, Tsutomu Tanaka, Hirofumi Ichinose, 神谷 典穂, Synergistic effect and application of xylanases as accessory enzymes to enhance the hydrolysis of pretreated bagasse, ENZYME AND MICROBIAL TECHNOLOGY, 10.1016/j.enzmictec.2015.01.007, 72, 16-24, 2015.05. |
| 171. |
Synergistic effect and application of xylanases as accessory enzymes to enhance the hydrolysis of pretreated bagasse.. |
| 172. |
Momoko Kitaoka, Ayaka Naritomi, Yuya Hirakawa, Noriho Kamiya, Masahiro Goto, Transdermal immunization using solid-in-oil nanodispersion with CpG oligodeoxynucleotide adjuvants, Pharmaceutical Research, 10.1007/s11095-014-1554-5, 32, 4, 1486-1492, 2015.04, Purpose: Simple and noninvasive vaccine administration alternatives to injections are desired. A solid-in-oil (S/O) nanodispersion system was able to overcome skin barriers and induce an immune response; however, antibody levels remained low. We applied an immune potentiator, CpG oligodeoxynucleotide (ODN), to enhance the immune response by controlling the T helper 1 (Th1)/T helper 2 (Th2) balance. Methods: S/O nanodispersions containing ovalbumin (OVA) and CpG ODN (CpG-A or CpG-B) were characterized by size distribution analysis and a protein release test. The skin permeation of fluorescence-labeled OVA was observed by fluorescence microscopy. Antigen-specific IgG, IgG1, and IgG2a responses were measured by enzyme-linked immunosorbent assay. Results: Co-encapsulation of CpG ODNs in S/O nanodispersions enhanced induction of OVA-specific IgG. S/O nanodispersion containing OVA and CpG-A had a smaller mean particle size and permeated the skin more efficiently. In contrast, CpG-B showed the highest protein release and induction of OVA-specific IgG. IgG subclass analysis revealed that OVA induced a Th2-dominant immune response, while the S/O nanodispersion containing CpG-A skewed the immune response toward a Th1-bias. Conclusions: In combination with CpG ODN, the S/O nanodispersion system efficiently induced an antigen-specific antibody response. The Th1/Th2 immune balance could be controlled by the selection of CpG ODN type.. |
| 173. |
Kousuke Moriyama, Rie Wakabayashi, Masahiro Goto, 神谷 典穂, Enzyme-mediated preparation of hydrogels composed of poly(ethylene glycol) and gelatin as cell culture platforms, RSC ADVANCES, 10.1039/c4ra12334d, 5, 4, 3070-3073, 2015.03. |
| 174. |
Kounosuke Hayashi, Yusuke Tomozoe, Kenji Nagai, Yoshiyuki Hiraishi, Noriho Kamiya, Development of a peroxidase-Fused protein reagent by brevibacillus choshinensis heterologous expression system, kagaku kogaku ronbunshu, 10.1252/kakoronbunshu.41.157, 41, 2, 157-161, 2015.03, To detect a target protein in biological samples, a fusion protein was designed composed of a peroxidase from Arthromyces ramosus (ARP) and parts of the antibody-binding domains of Staphylococcus aureus protein A and Streptococcus protein G (PG). The ARP-PG fusion protein was successfully expressed by a heterologous protein expression system in Brevibacillus choshinensis. The fusion protein was secreted as an active form in culture media. The production of ARP-PG with higher peroxidase activity was observed by the addition of 5-aminolevulinic acid to the culture media. The performance of purified ARP-PG was validated by dot blotting for the detection of transferrin as a model target protein. A comparable performance in the dot blot analysis was attained using a culture supernatant containing crude but active ARP-PG, indicating the practicality of the Brevibacillus protein secretion system.. |
| 175. |
Jian Yang, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Separation of gold(III) in acidic chloride solution using porous polymeric ionic liquid gel, JOURNAL OF CHEMICAL ENGINEERING OF JAPAN, 10.1252/jcej.14we162, 48, 3, 197-201, 2015.03, A novel polymeric ionic liquid (PIL) gel was synthesized via radical polymerization of the ionic liquid monomers, 1-vinyl- 3-ethylimidazoliumbis(trifluorometyl-sulfonyl) imide ([veim][Tf2N]). The gel was made porous by removing the water phase dispersed by polyoxyethylene (20) sorbitan monolaurate (Tween-20). The porous structure of the synthesized gel was observed by SEM imaging. In comparison to the nonporous PIL gel synthesized without Tween-20, the adsorption capacity of Au(III) onto porous PIL gel was enhanced. The adsorption behavior of the synthesized gel was studied in acidic chloride solutions containing various metal ions, and it was found that the gel selectively adsorbed Au(III) over other metal ions. Maximum adsorption capacity of Au(III) was more than 140 mg/g; whereas, those for Pt(IV) and Pd(II) were lower than 20 mg/g. The adsorbed Au(III) onto the gel can easily be desorbed using an acidified thiourea aqueous solution. The feasibility of utilizing the polymeric ionic liquid gel for the separation of Au(III) was thereby demonstrated.. |
| 176. |
Katsuyuki Miyawaki, Sumihare Noji, Noriho Kamiya, Transglutaminase-mediated in situ hybridization (transish) for mRNA detection in mammalian tissues, In Situ Hybridization Methods, 10.1007/978-1-4939-2303-8_29, 549-558, 2015.02, In the most commonly used in situ hybridization (ISH) procedure, a hapten-labeled antisense nucleic acid (e.g., RNA) probe is employed to hybridize a target mRNA in tissue sections. The hapten-labeled RNA is then detected by a highly specific hapten-antibody interaction; however, it requires laborious immunostaining steps for spatial coloring on tissue sections. To simplify ISH-based mRNA detection systems, we created a new RNA-(enzyme) n conjugate for sensitive detection of mRNA in tissue sections. In the present simple, antibody-free ISH protocol, an antisense RNA probe of interest is first modified with a specific dipeptide substrate of microbial transglutaminase (MTG) by in vitro transcription. Alkaline phosphatase from a hyperthermophile is then covalently linked to the substrate-labeled RNA by MTG-catalyzed sitespecific conjugation. A robust, multi-enzyme-labeled RNA probe enables the direct labeling of a target mRNA in tissue sections with signaling enzymes under harsh hybridization conditions, leading to one-step signal amplification after hybridization. The application of the new trans glutaminase-mediated ISH (TransISH) strategy to mRNA detection in mammalian tissues was demonstrated.. |
| 177. |
Katsuyuki Miyawaki, Sumihare Noji, Noriho Kamiya, Transglutaminase-mediated in situ hybridization (transish) for mRNA detection in mammalian tissues, In Situ Hybridization Methods, 10.1007/978-1-4939-2303-8_29, 549-558, 2015.02, In the most commonly used in situ hybridization (ISH) procedure, a hapten-labeled antisense nucleic acid (e.g., RNA) probe is employed to hybridize a target mRNA in tissue sections. The hapten-labeled RNA is then detected by a highly specific hapten-antibody interaction
however, it requires laborious immunostaining steps for spatial coloring on tissue sections. To simplify ISH-based mRNA detection systems, we created a new RNA-(enzyme) n conjugate for sensitive detection of mRNA in tissue sections. In the present simple, antibody-free ISH protocol, an antisense RNA probe of interest is first modified with a specific dipeptide substrate of microbial transglutaminase (MTG) by in vitro transcription. Alkaline phosphatase from a hyperthermophile is then covalently linked to the substrate-labeled RNA by MTG-catalyzed sitespecific conjugation. A robust, multi-enzyme-labeled RNA probe enables the direct labeling of a target mRNA in tissue sections with signaling enzymes under harsh hybridization conditions, leading to one-step signal amplification after hybridization. The application of the new trans glutaminase-mediated ISH (TransISH) strategy to mRNA detection in mammalian tissues was demonstrated.. |
| 178. |
Kousuke Moriyama, Rie Wakabayashi, Masahiro Goto, 神谷 典穂, Characterization of Enzymatically Gellable, Phenolated Linear Poly(Ethylene Glycol) with Different Molecular Weights for Encapsulating Living Cells, BIOCHEMICAL ENGINEERING JOURNAL, 93, 25-30, 2015.01. |
| 179. |
Kosuke Minamihata, Masahiro Goto, 神谷 典穂, Site-specific conjugation of an antibody-binding protein catalyzed by horseradish peroxidase creates a multivalent protein conjugate with high affinity to IgG, BIOTECHNOLOGY JOURNAL, 10.1002/biot.201400512, 10, 1, 222-226, 2015.01. |
| 180. |
Siti Norulhuda Hashim, Akira Tsuchiya, Noriho Kamiya, Shinsuke Sando, A single fluorophore-labeled aptamer sensor for the detection of interferon gamma, Chemistry Letters, 10.1246/cl.150794, 44, 12, 1670-1672, 2015.01, Interferon gamma (IFN-γ) is a cytokine that is mainly secreted from immune cells. It plays an important role in regulating neighboring cellular activities through interactions with cell membrane proteins. Because of its biological importance, there has been increasing demand for a biosensor that can respond to IFN-γ. Here, we report a fluorescent aptamer sensor that targets IFN-γ. The 25-nt IFN-γ aptamer was converted into a turn-on fluorescent sensor by labeling the 3'-end with fluorescein. The designed sensor achieved fluorescence sensing of IFN-γ with nanomolar affinity.. |
| 181. |
M. Moniruzzaman, H. Mahmood, Noriho Kamiya, S. Yusup, Masahiro Goto, Activity and stability of enzyme immobilized with ionic liquid based polymer materials, Journal of Engineering Science and Technology, 10, 60-69, 2015.01, Enzyme immobilization methods are continuously being developed for a wide range of applications including biocatalysis and biotransformation. Compared to other immobilization methods, physical entrapment into a matrix is relatively simple and inexpensive, and causes a relatively small perturbation to the native enzyme structure and function. However, significant enzyme leaching from such biocatalytic polymers is the main constraint in using them for industrial levels. This study reports an ionic liquid (IL) polymer materials incorporating enzymes that can be used as active, stable and reusable biocatalysts to overcome the limitations. Lipase was microencapsulated in surfactant aggregates formed in an IL monomer or the solution of an IL monomer/IL and then incorporated into polymer frameworks through the free radical polymerization of an IL (1-vinyl-3-ethylimidazolium bis(trifluoromethylsulfonyl) amide ([veim][Tf2N]). The activity, stability and reusability of IL polymer materials containing lipase were evaluated using lipase-catalyzed hydrolysis of p-nitrophenyl butyrate (p-PNB) as a model reaction. It was found that lipase IL polymer materials exhibited excellent stability in aqueous solutions. More importantly, these biopolymer materials retained most of their activity after six reaction cycles. We firmly believe that enzyme containing IL polymeric materials developed in this study will provide several advantages over conventional polymer based methods for diverse enzymatic reactions.. |
| 182. |
Kounosuke Hayashi, Yusuke Tomozoe, Kenji Nagai, Kyoichi Matsuba, Masayuki Mitsumori, Yoshiyuki Hiraishi, Toshiharu Matsumura, Hiroshi Ueda, Noriho Kamiya, Optimization of a fusion protein expression system using human cell lines to create a practical immunoassay reagent, kagaku kogaku ronbunshu, 10.1252/kakoronbunshu.41.38, 41, 1, 38-42, 2015.01, New fusion proteins for immunoassay were developed as an alternative to the conventional chemical linked enzyme-antibody complex. Human chimeric alkaline phosphatase (IPP) was used as the labeling enzyme, and partial domains of Staphylococcus aureus Protein A and Streptococcus Protein G were used as antibody binding protein (PG). The fusion protein composed of IPP and PG was produced using human cell lines because IPP is derived from human enzymes. The expression system was optimized by changing the plasmid vector, reducing the GC content of the DNA sequence, and employing different cell lines including adherent and suspension cells. As result, a 2,600-fold increase in the protein yield per unit of growth medium was achieved. The resultant IPP-PG fusion protein was successfully utilized for immunoassay applications such as western blotting and immunohistochemistry.. |
| 183. |
Katsuyuki Miyawaki, Sumihare Noji, Noriho Kamiya, Transglutaminase-mediated in situ hybridization (TransISH) for mRNA detection in mammalian tissues, Neuromethods, 10.1007/978-1-4939-2303-8_29, 99, 549-558, 2015.01, In the most commonly used in situ hybridization (ISH) procedure, a hapten-labeled antisense nucleic acid (e.g., RNA) probe is employed to hybridize a target mRNA in tissue sections. The hapten-labeled RNA is then detected by a highly specific hapten–antibody interaction; however, it requires laborious immunostaining steps for spatial coloring on tissue sections. To simplify ISH-based mRNA detection systems, we created a new RNA–(enzyme) n conjugate for sensitive detection of mRNA in tissue sections. In the present simple, antibody-free ISH protocol, an antisense RNA probe of interest is first modified with a specific dipeptide substrate of microbial transglutaminase (MTG) by in vitro transcription. Alkaline phosphatase from a hyperthermophile is then covalently linked to the substrate-labeled RNA by MTG-catalyzed sitespecific conjugation. A robust, multi-enzyme-labeled RNA probe enables the direct labeling of a target mRNA in tissue sections with signaling enzymes under harsh hybridization conditions, leading to one-step signal amplification after hybridization. The application of the new trans glutaminase-mediated ISH (TransISH) strategy to mRNA detection in mammalian tissues was demonstrated.. |
| 184. |
Activity and stability of enzyme immobilized with ionic liquid based polymer materials
© School of Engineering, Taylor’s University. Enzyme immobilization methods are continuously being developed for a wide range of applications including biocatalysis and biotransformation. Compared to other immobilization methods, physical entrapment into a matrix is relatively simple and inexpensive, and causes a relatively small perturbation to the native enzyme structure and function. However, significant enzyme leaching from such biocatalytic polymers is the main constraint in using them for industrial levels. This study reports an ionic liquid (IL) polymer materials incorporating enzymes that can be used as active, stable and reusable biocatalysts to overcome the limitations. Lipase was microencapsulated in surfactant aggregates formed in an IL monomer or the solution of an IL monomer/IL and then incorporated into polymer frameworks through the free radical polymerization of an IL (1-vinyl-3-ethylimidazolium bis(trifluoromethylsulfonyl) amide ([veim][Tf2N]). The activity, stability and reusability of IL polymer materials containing lipase were evaluated using lipase-catalyzed hydrolysis of p-nitrophenyl butyrate (p-PNB) as a model reaction. It was found that lipase IL polymer materials exhibited excellent stability in aqueous solutions. More importantly, these biopolymer materials retained most of their activity after six reaction cycles. We firmly believe that enzyme containing IL polymeric materials developed in this study will provide several advantages over conventional polymer based methods for diverse enzymatic reactions.. |
| 185. |
Enzyme-mediated preparation of hydrogels composed of poly(ethylene glycol) and gelatin as cell culture platforms
© The Royal Society of Chemistry 2015. A redox-responsive hydrogel composed of poly(ethylene glycol) and gelatin was prepared via an enzymatic oxidative reaction. Fibroblast cells adhered on the hydrogel showed proliferation and they could be recovered as a cell sheet by degradation of the scaffold under mild reductive conditions, thus providing a novel platform for cell culture.. |
| 186. |
Ionic liquid-mediated transcutaneous protein delivery with solid-in-oil nanodispersions
© 2015 The Royal Society of Chemistry. As a potentially safe and non-invasive vaccination method, transcutaneous immunization represents an attractive alternative to conventional vaccine delivery by injection. However, the development of transcutaneous immunization has remained a challenge for a large number of hydrophilic macromolecules including protein and peptide antigens. We report a novel ionic liquid (IL)-mediated transcutaneous vaccine formulation consisting of a solid-in-oil (S/O) nanodispersion of antigen coated with pharmaceutically accepted surfactants dispersed in IL-containing oil. The introduction of the IL [C12mim][Tf2N] (1-dodecyl-3-methyl imidazolium bis(trifluoromethyl sulfonyl) amide) as a penetration enhancer in the formulation significantly enhanced the skin permeability of ovalbumin (OVA), a model antigen. It was also found that the IL-mediated S/O nanodispersion obtained high levels of OVA-specific serum IgG compared with both S/O nanodispersions without IL and PBS control. These findings clearly indicate that ILs-which are potentially attractive "green" and "designer" solvents-could serve as potential skin penetration enhancers in transcutaneous vaccination for hydrophilic macromolecules.. |
| 187. |
Separation of gold(III) in acidic chloride solution using porous polymeric ionic liquid gel
© 2015 The Society of Chemical Engineers, Japan. A novel polymeric ionic liquid (PIL) gel was synthesized via radical polymerization of the ionic liquid monomers, 1-vinyl- 3-ethylimidazoliumbis(trifluorometyl-sulfonyl) imide ([veim][Tf2N]). The gel was made porous by removing the water phase dispersed by polyoxyethylene (20) sorbitan monolaurate (Tween-20). The porous structure of the synthesized gel was observed by SEM imaging. In comparison to the nonporous PIL gel synthesized without Tween-20, the adsorption capacity of Au(III) onto porous PIL gel was enhanced. The adsorption behavior of the synthesized gel was studied in acidic chloride solutions containing various metal ions, and it was found that the gel selectively adsorbed Au(III) over other metal ions. Maximum adsorption capacity of Au(III) was more than 140 mg/g; whereas, those for Pt(IV) and Pd(II) were lower than 20 mg/g. The adsorbed Au(III) onto the gel can easily be desorbed using an acidified thiourea aqueous solution. The feasibility of utilizing the polymeric ionic liquid gel for the separation of Au(III) was thereby demonstrated.. |
| 188. |
Site-specific conjugation of an antibody-binding protein catalyzed by horseradish peroxidase creates a multivalent protein conjugate with high affinity to IgG
© 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim. Cross-linking proteins offers an approach to enhance the distinct function of proteins due to the multivalent effect. In this study, we demonstrated the preparation of a multivalent antibody-binding protein possessing high affinity to IgG by conjugating a number of antibody-binding proteins using the horseradish peroxidase (HRP)-mediated protein conjugation method. By introducing a peptide tag containing a tyrosine (Y-tag) to the C-terminus of the model protein, a chimera protein of protein G and protein A (pG2pA), the Tyr residue in the Y-tag was efficiently recognized by HRP and cross-linked with each other to yield a pG2pA conjugate, composed of mainly two to three units of pG2pA. The cross-linking occurred site specifically at the Tyr residue in the Y-tag and introduction of the Y-tag showed no effect on the function of pG2pA. The affinity of the Y-tagged pG2pA conjugate against IgG clearly increased because of the multivalent effect, demonstrating the benefit of this protein cross-linking reaction, which yields functional protein oligomers. Such multivalent protein conjugates created by this reaction should have potential to be used in ELISA and Western blotting applications in which highly sensitive detection of target molecules is desired.. |
| 189. |
Transdermal immunization using solid-in-oil nanodispersion with CpG oligodeoxynucleotide adjuvants
© 2014 Springer Science+Business Media New York. Purpose: Simple and noninvasive vaccine administration alternatives to injections are desired. A solid-in-oil (S/O) nanodispersion system was able to overcome skin barriers and induce an immune response; however, antibody levels remained low. We applied an immune potentiator, CpG oligodeoxynucleotide (ODN), to enhance the immune response by controlling the T helper 1 (Th1)/T helper 2 (Th2) balance. Methods: S/O nanodispersions containing ovalbumin (OVA) and CpG ODN (CpG-A or CpG-B) were characterized by size distribution analysis and a protein release test. The skin permeation of fluorescence-labeled OVA was observed by fluorescence microscopy. Antigen-specific IgG, IgG1, and IgG2a responses were measured by enzyme-linked immunosorbent assay. Results: Co-encapsulation of CpG ODNs in S/O nanodispersions enhanced induction of OVA-specific IgG. S/O nanodispersion containing OVA and CpG-A had a smaller mean particle size and permeated the skin more efficiently. In contrast, CpG-B showed the highest protein release and induction of OVA-specific IgG. IgG subclass analysis revealed that OVA induced a Th2-dominant immune response, while the S/O nanodispersion containing CpG-A skewed the immune response toward a Th1-bias. Conclusions: In combination with CpG ODN, the S/O nanodispersion system efficiently induced an antigen-specific antibody response. The Th1/Th2 immune balance could be controlled by the selection of CpG ODN type.. |
| 190. |
Shota Araki, Rie Wakabayashi, Muhammad Moniruzzaman, Noriho Kamiya, Masahiro Goto, Ionic liquid-mediated transcutaneous protein delivery with solid-in-oil nanodispersions, MedChemComm, 10.1039/c5md00378d, 6, 12, 2124-2128, 2015, As a potentially safe and non-invasive vaccination method, transcutaneous immunization represents an attractive alternative to conventional vaccine delivery by injection. However, the development of transcutaneous immunization has remained a challenge for a large number of hydrophilic macromolecules including protein and peptide antigens. We report a novel ionic liquid (IL)-mediated transcutaneous vaccine formulation consisting of a solid-in-oil (S/O) nanodispersion of antigen coated with pharmaceutically accepted surfactants dispersed in IL-containing oil. The introduction of the IL [C12mim][Tf2N] (1-dodecyl-3-methyl imidazolium bis(trifluoromethyl sulfonyl) amide) as a penetration enhancer in the formulation significantly enhanced the skin permeability of ovalbumin (OVA), a model antigen. It was also found that the IL-mediated S/O nanodispersion obtained high levels of OVA-specific serum IgG compared with both S/O nanodispersions without IL and PBS control. These findings clearly indicate that ILs-which are potentially attractive "green" and "designer" solvents-could serve as potential skin penetration enhancers in transcutaneous vaccination for hydrophilic macromolecules.. |
| 191. |
Rie Wakabayashi, Ryutaro Ishiyama, Noriho Kamiya, Masahiro Goto, A novel surface-coated nanocarrier for efficient encapsulation and delivery of camptothecin to cells, MedChemComm, 10.1039/c4md00179f, 5, 10, 1515-1519, 2014.10, In the present study, we developed a novel surface-coated nanocarrier (SCN) for efficient and stable encapsulation of a poorly water-soluble anticancer agent, camptothecin (CPT). Using emulsification and freeze-drying processes in the presence of a hydrophilic surfactant, Pluronic F-68 or F-127, CPT-Pluronic complexes with crystalline features were synthesised. Investigation of the in vitro anticancer activities and cellular internalisation of the complexes revealed that the SCN efficiently delivered CPT to cells while maintaining anticancer activity.. |
| 192. |
Jian Yang, Fukiko Kubota, Yuzo Baba, Noriho Kamiya, Masahiro Goto, Application of cellulose acetate to the selective adsorption and recovery of Au(III), Carbohydrate Polymers, 10.1016/j.carbpol.2014.05.003, 111, 768-774, 2014.10, Cellulose acetyl derivatives were examined for the selective recovery of Au(III) from acidic chloride solutions as an adsorbent, and cellulose acetate fibers (CAF) were found to be effective for the separation of Au(III) from other metal ions, including the precious metal ions Pt(IV) and Pd(II). The amount of Au(III) adsorbed by the fibers increased with an increase in the hydrochloric acid concentration, but decreased with an increase in the ionic strength of the solution. The adsorption of Au(III) onto CAF took place quickly and an adsorption equilibrium was reached within 1 h. The maximum adsorption capacity of Au(III) was determined to be 110 mg/g at 2 M hydrochloric acid. The loaded Au(III) was readily recovered by incineration.. |
| 193. |
A novel surface-coated nanocarrier for efficient encapsulation and delivery of camptothecin to cells
© 2014 the Partner Organisations. In the present study, we developed a novel surface-coated nanocarrier (SCN) for efficient and stable encapsulation of a poorly water-soluble anticancer agent, camptothecin (CPT). Using emulsification and freeze-drying processes in the presence of a hydrophilic surfactant, Pluronic F-68 or F-127, CPT-Pluronic complexes with crystalline features were synthesised. Investigation of the in vitro anticancer activities and cellular internalisation of the complexes revealed that the SCN efficiently delivered CPT to cells while maintaining anticancer activity.. |
| 194. |
Application of cellulose acetate to the selective adsorption and recovery of Au(III)
Cellulose acetyl derivatives were examined for the selective recovery of Au(III) from acidic chloride solutions as an adsorbent, and cellulose acetate fibers (CAF) were found to be effective for the separation of Au(III) from other metal ions, including the precious metal ions Pt(IV) and Pd(II). The amount of Au(III) adsorbed by the fibers increased with an increase in the hydrochloric acid concentration, but decreased with an increase in the ionic strength of the solution. The adsorption of Au(III) onto CAF took place quickly and an adsorption equilibrium was reached within 1 h. The maximum adsorption capacity of Au(III) was determined to be 110 mg/g at 2 M hydrochloric acid. The loaded Au(III) was readily recovered by incineration. © 2014 Elsevier Ltd.. |
| 195. |
Kousuke Moriyama, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya, Characterization of enzymatically gellable, phenolated linear poly(ethylene glycol) with different molecular weights for encapsulating living cells, Biochemical Engineering Journal, 10.1016/j.bej.2014.09.003, 93, 25-30, 2014.09, Enzymatic hydrogelation has received much attention due to the high biocompatibility and the ease of control of reaction kinetics under physiological conditions. In particular, horseradish peroxidase (HRP)-mediated phenol coupling reaction has great potential for developing in situ hydrogelation systems. Herein, we report the HRP-catalyzed preparation and characterization of hydrogels composed of a terminally bis-phenolated linear poly(ethylene glycol) (PEG-Ph-OH) with different molecular weights (Mws 3100, 8800, 11,000, 20,000g/mol). The gelation time of polymer solution can be controlled in the range from few second to few minute, suggestion that the PEG-Ph-OH has a potential as a in situ forming hydrogel. In addition, the physicochemical properties of the hydrogels, such as swelling ratio, mesh size and mechanical property, were controlled by the molecular weight of the PEG-Ph-OH. The results could be attributed to the alteration in the cross-linking density by the variation of molecular weight of the gel precursor. Furthermore, the viability of mammalian cells encapsulated in the PEG-Ph-OH hydrogels was approximately 90%. These results indicate that PEG-Ph-OH has potential for biomedical applications including tissue engineering.. |
| 196. |
Characterization of enzymatically gellable, phenolated linear poly(ethylene glycol) with different molecular weights for encapsulating living cells
© 2014 Elsevier B.V. Enzymatic hydrogelation has received much attention due to the high biocompatibility and the ease of control of reaction kinetics under physiological conditions. In particular, horseradish peroxidase (HRP)-mediated phenol coupling reaction has great potential for developing in situ hydrogelation systems. Herein, we report the HRP-catalyzed preparation and characterization of hydrogels composed of a terminally bis-phenolated linear poly(ethylene glycol) (PEG-Ph-OH) with different molecular weights (Mws 3100, 8800, 11,000, 20,000g/mol). The gelation time of polymer solution can be controlled in the range from few second to few minute, suggestion that the PEG-Ph-OH has a potential as a in situ forming hydrogel. In addition, the physicochemical properties of the hydrogels, such as swelling ratio, mesh size and mechanical property, were controlled by the molecular weight of the PEG-Ph-OH. The results could be attributed to the alteration in the cross-linking density by the variation of molecular weight of the gel precursor. Furthermore, the viability of mammalian cells encapsulated in the PEG-Ph-OH hydrogels was approximately 90%. These results indicate that PEG-Ph-OH has potential for biomedical applications including tissue engineering.. |
| 197. |
Kousuke Moriyama, Kosuke Minamihata, Rie Wakabayashi, Masahiro Goto, 神谷 典穂, Enzymatic preparation of a redox-responsive hydrogel for encapsulating and releasing living cells, CHEMICAL COMMUNICATIONS, 10.1039/c3cc49766f, 50, 44, 5895-5898, 2014.08, Horseradish peroxidase-mediated oxidative cross-linking of a thiolated poly(ethylene glycol) is promoted in the absence of exogenous hydrogen peroxide, by adding a small amount of phenolic compound under physiological conditions. The prepared hydrogel can encapsulate and release living mammalian cells.. |
| 198. |
Zhigang Zhao, Yuzo Baba, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Synergistic Extraction of Rare-Earth Metals and Separation of Scandium Using 2-Thenoyltriuoroacetone and Tri-n-octylphosphine Oxide in an Ionic Liquid System, JOURNAL OF CHEMICAL ENGINEERING OF JAPAN, 10.1252/jcej.14we360, 47, 8, 656-662, 2014.08, The synergistic extraction of rare-earth metals by a beta-diketone and an organophosphine oxide is described. A combination of 2-thenoyltri'uor oacetone (HTTA) and tri-n-octyl phosphine oxide (TOPO) was found to be e degrees ective in synergistically extracting rare-earth metals into an ionic liquid (IL). The synergistic e degrees ect was strongly in(similar to)uenc ed by the diluent in which the extractants were dissolved. Extraction e ciencies for all the rare-earth metal ions were enhanced by synergy when a conventional organic solvent was used (n-dodecane). However, the synergistic e degrees ect in the IL preferentially improved the scandium extraction, leading to higher selectivity for Sc than for other rare-earth metals. Slope analysis was used to assess each extraction system, and the IL itself was found to be directly involved in the complex formation of the metal extraction system.. |
| 199. |
Rie Wakabayashi, Yuko Abe, Masahiro Goto, 神谷 典穂, The Self-Assembly and Secondary Structure of Peptide Amphiphiles Determine the Membrane Permeation Activity, RSC Advances, 4, 30654-30657, 2014.07. |
| 200. |
Rie Wakabayashi, Ryutaro Ishiyama, 神谷 典穂, Masahiro Goto, A Novel Surface-Coated Nanocarrier for Efficient Encapsulation and Delivery of Camptothecin to Cells, MedChemComm, 5, 1515-1519, 2014.07. |
| 201. |
Teppei Niide, Masahiro Goto, 神谷 典穂, Enzymatic self-sacrificial display of an active protein on gold nanoparticles, RSC ADVANCES, 10.1039/c3ra46384b, 4, 12, 5995-5998, 2014.04. |
| 202. |
Baba, Yuzo, Fukiko Kubota, 神谷 典穂, Masahiro Goto, Development of Novel Extractants with Amino Acid Structure for Efficient Separation of Nickel and Cobalt from Manganese Ions, INDUSTRIAL & ENGINEERING CHEMISTRY RESEARCH, 10.1021/ie403524a, 53, 2, 812-818, 2014.01. |
| 203. |
Kitaoka, Momoko, Imamura, Kana, Hirakawa, Yuya, Tahara, Yoshiro, 神谷 典穂, Masahiro Goto, Sucrose laurate-enhanced transcutaneous immunization with a solid-in-oil nanodispersion, MEDCHEMCOMM, 10.1039/c3md00164d, 5, 1, 20-24, 2014.01. |
| 204. |
Jameson, Laramie P., Balaz, Milan, Dzyuba, Sergei V., 神谷 典穂, Conformational preference of a porphyrin rotor in confined environments, RSC ADVANCES, 10.1039/c3ra45668d, 4, 2, 705-708, 2014.01. |
| 205. |
Yuzo Baba, Arisa Fukami, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Selective extraction of scandium from yttrium and lanthanides with amic acid-type extractant containing alkylamide and glycine moieties, RSC Advances, 10.1039/c4ra08897b, 4, 92, 50726-50730, 2014.01, The liquid-liquid extraction of rare earth metal ions (scandium (Sc3+), yttrium (Y3+) and the lanthanides (La3+, Nd3+, Eu3+ and Dy3+)) was investigated using N-[N,N-di(2-ethylhexyl)aminocarbonylmethyl]glycine (D2EHAG). Scandium was extracted selectively from lanthanides under highly acidic conditions (0 -3 H2SO4. By comparing the extraction behavior with N,N-dioctyldiglycol amic acid, which has a similar molecular structure to D2EHAG, or the commercial alkyl monocarboxylic acid extractant, Versatic 10, it was concluded that the affinity of D2EHAG to scandium was caused by a chelating effect and the size recognition ability of D2EHAG. The extraction mechanism was examined, and it was proven that the trivalent scandium ion is extracted by forming a stable metal complex with four D2EHAG molecules. This journal is. |
| 206. |
Development of novel extractants with amino acid structure for efficient separation of nickel and cobalt from manganese ions
The solvent extraction separation of nickel and cobalt from manganese is an important issue to be resolved. Novel amic acid type extractants with the glycine or sarcosine moiety N-[N,N-di(2-ethylhexyl)aminocarbonylmethyl]glycine (D2EHAG) and N-[N,N-di(2-ethylhexyl)aminocarbonylmethyl]sarcosine (D2EHAS) were synthesized to achieve this separation. These extractants were prepared by a simple two-step reaction and were readily soluble in aliphatic organic solvents. The extraction behavior of Ni2+, Co2+, and Mn 2+ with D2EHAG and D2EHAS in n-dodecane was investigated compared with that of conventional extractants. The novel extractants extracted Ni 2+ and Co2+ preferentially with a high selectivity to Mn2+. The metal ions could be back-extracted quantitatively using 1 mol dm-3 sulfuric acid. The extraction mechanism of the three metal ions with D2EHAG and D2EHAS was investigated, and results suggest that one divalent metal ion is extracted with two extractant molecules. © 2013 American Chemical Society.. |
| 207. |
One step effective separation of platinum and palladium in an acidic chloride solution by using undiluted ionic liquids
Imidazolium-based ionic liquids (ILs) with Tf2N- as anions were studied as extractants without dilution for the extractive separation of platinum and palladium in an acidic chloride solution. The effect of the alkyl chain length on the extraction performance was evaluated and, based on the performance, the IL, 1-methyl-3-octylimidazolium bis(trifluoromethylsulfonyl) imide ([Omim][Tf2N]), was selected to be studied in detail. The extraction of Pt(IV) underwent little change over the HCl concentration studied, while that of Pd(II) decreased with increasing HCl concentration. The separation of Pt(IV) and Pd(II) was highly effective, and the highest separation factor between Pt(IV) and Pd(II) reached 312 at the optimum conditions in a binary component system composed of Pt(IV) and Pd(II). The effect of the initial Pt(IV) concentration on loading capacity and extraction efficiency of Pt(IV) was also investigated.. |
| 208. |
Separation of precious metals by using undiluted ionic liquids
The ionic liquid (IL), 1-butyl-3-methylimidazolium bis(trifluoromethylsulfonyl)imide ([Bmim][Tf2N]), was evaluated as an extractant without dilution for the separation and recovery of precious metals. Au(III) was efficiently extracted from the aqueous phase to the IL phase, and a concentrated Au(III) solution almost 8 times higher than the initial feed solutions could be produced. In contrast, Pd(II) was rarely extracted and Pt(IV) was partly extracted under the present experimental conditions. Furthermore, the effect of the IL anions on the extraction ability for the metal ions was examined, and it was found that an IL having an anion component slightly more hydrophilic compared to [Tf2N], such as 1-butyl-3-methylimidazolium hexafluorophosphate [Bmim][PF6], showed a higher extraction ability than [Bmim][Tf2N]. It was suggested that the extraction proceeded via an anion exchange mechanism and it was shown that ILs could be effective extractants for the separation of precious metals.. |
| 209. |
Sucrose laurate enhanced transcutaneous immunization with a solid-in-oil nanodispersion. |
| 210. |
The self-assembly and secondary structure of peptide amphiphiles determine the membrane permeation activity
Membrane fusogenic peptides have attracted increasing attention because of their unique biofunctions in membrane translocation and viral infection. Here, we designed GALA-related peptides with palmitoyl tails. Our study indicated that the self-assembling propensity and the secondary structure of these peptide amphiphiles greatly influenced the membrane permeability. This journal is © the Partner Organisations 2014.. |
| 211. |
Jian Yang, Fukiko Kubota, Yuzo Baba, Noriho Kamiya, Masahiro Goto, One step effective separation of platinum and palladium in an acidic chloride solution by using undiluted ionic liquids, Solvent Extraction Research and Development, 10.15261/serdj.21.129, 21, 2, 129-135, 2014, Imidazolium-based ionic liquids (ILs) with Tf2N- as anions were studied as extractants without dilution for the extractive separation of platinum and palladium in an acidic chloride solution. The effect of the alkyl chain length on the extraction performance was evaluated and, based on the performance, the IL, 1-methyl-3-octylimidazolium bis(trifluoromethylsulfonyl) imide ([Omim][Tf2N]), was selected to be studied in detail. The extraction of Pt(IV) underwent little change over the HCl concentration studied, while that of Pd(II) decreased with increasing HCl concentration. The separation of Pt(IV) and Pd(II) was highly effective, and the highest separation factor between Pt(IV) and Pd(II) reached 312 at the optimum conditions in a binary component system composed of Pt(IV) and Pd(II). The effect of the initial Pt(IV) concentration on loading capacity and extraction efficiency of Pt(IV) was also investigated.. |
| 212. |
Jian Yang, Fukiko Kubota, Yuzo Baba, Noriho Kamiya, Masahiro Goto, Separation of precious metals by using undiluted ionic liquids, Solvent Extraction Research and Development, 10.15261/serdj.21.89, 21, 1, 89-94, 2014, The ionic liquid (IL), 1-butyl-3-methylimidazolium bis(trifluoromethylsulfonyl)imide ([Bmim][Tf2N]), was evaluated as an extractant without dilution for the separation and recovery of precious metals. Au(III) was efficiently extracted from the aqueous phase to the IL phase, and a concentrated Au(III) solution almost 8 times higher than the initial feed solutions could be produced. In contrast, Pd(II) was rarely extracted and Pt(IV) was partly extracted under the present experimental conditions. Furthermore, the effect of the IL anions on the extraction ability for the metal ions was examined, and it was found that an IL having an anion component slightly more hydrophilic compared to [Tf2N], such as 1-butyl-3-methylimidazolium hexafluorophosphate [Bmim][PF6], showed a higher extraction ability than [Bmim][Tf2N]. It was suggested that the extraction proceeded via an anion exchange mechanism and it was shown that ILs could be effective extractants for the separation of precious metals.. |
| 213. |
Zhigang Zhao, Yuzo Baba, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Noriho Kamiya, Masahiro Goto, Synergistic extraction of rare-earth metals and separation of scandium using 2-thenoyltrifluoroacetone and tri-n-octylphosphine oxide in an ionic liquid system, JOURNAL OF CHEMICAL ENGINEERING OF JAPAN, 10.1252/jcej.14we360, 47, 8, 656-662, 2014, The synergistic extraction of rare-earth metals by a β-diketone and an organophosphine oxide is described. A combination of 2-thenoyltrifluoroacetone (HTTA) and tri-n-octyl phosphine oxide (TOPO) was found to be effective in synergistically extracting rare-earth metals into an ionic liquid (IL). The synergistic effect was strongly influenced by the diluent in which the extractants were dissolved. Extraction efficiencies for all the rare-earth metal ions were enhanced by synergy when a conventional organic solvent was used (n-dodecane). However, the synergistic effect in the IL preferentially improved the scandium extraction, leading to higher selectivity for Sc than for other rare-earth metals. Slope analysis was used to assess each extraction system, and the IL itself was found to be directly involved in the complex formation of the metal extraction system.. |
| 214. |
Rie Wakabayashi, Yuko Abe, Noriho Kamiya, Masahiro Goto, The self-assembly and secondary structure of peptide amphiphiles determine the membrane permeation activity, RSC Advances, 10.1039/c4ra02901a, 4, 58, 30654-30657, 2014, Membrane fusogenic peptides have attracted increasing attention because of their unique biofunctions in membrane translocation and viral infection. Here, we designed GALA-related peptides with palmitoyl tails. Our study indicated that the self-assembling propensity and the secondary structure of these peptide amphiphiles greatly influenced the membrane permeability. This journal is. |
| 215. |
Takahara, Mari, Hayashi, Kounosuke, Masahiro Goto, 神谷 典穂, Tailing DNA aptamers with a functional protein by two-step enzymatic reaction, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 10.1016/j.jbiosc.2013.05.025, 116, 6, 660-665, 2013.12. |
| 216. |
Niide, Teppei, Shimojo, Kojiro, Rie Wakabayashi, Masahiro Goto, 神谷 典穂, Enzymatic Fabrication of Protein-Decorated Gold Nanoparticles by the Aid of Artificial Peptides with Gold-Binding Affinity, LANGMUIR, 10.1021/la401327h, 29, 50, 15596-15605, 2013.12. |
| 217. |
Kitaoka, Momoko, Imamura, Kana, Hirakawa, Yuya, Tahara, Yoshiro, 神谷 典穂, Masahiro Goto, Needle-free immunization using a solid-in-oil nanodispersion enhanced by a skin-permeable oligoarginine peptide, INTERNATIONAL JOURNAL OF PHARMACEUTICS, 10.1016/j.ijpharm.2013.10.006, 458, 2, 334-339, 2013.12. |
| 218. |
Enzymatic fabrication of protein-decorated gold nanoparticles by the aid of artificial peptides with gold-binding affinity
Here, we report a new approach for the biofabrication of protein-immobilized gold nanoparticles (Au NPs), using oxidoreductase with gold-binding peptide-tagged recombinant proteins. The reduction of Au ions to Au0 was achieved using a natural electron-donating cofactor, nicotinamide adenine dinucleotide, which was regenerated by the glycerol dehydrogenase (GLD) enzyme. First, we selected the A3 peptide (AYSSGAPPMPPF) as a gold binding moiety. The A3 peptide was introduced to the C-terminus of fusion proteins of immunoglobulin G (IgG)-binding domains of protein G and protein A. In the presence of the recombinant protein, the GLD-catalyzed cofactor reduction resulted in the efficient in situ fabrication of Au NPs immobilized with the fusion protein. Moreover, the protein-immobilized Au NPs were shown to have IgG binding activity. Although the A3 peptide had the ability to stabilize Au NPs, the results suggested that its binding affinity for Au NPs was unexpectedly weaker than that of His-tag. A cysteine residue was thus introduced to a recombinant protein adjacent to the A3 peptide. Finally, an artificial peptide, comprising A3 sequence with the C-terminal single cysteine residue, enabled the stable display of a fusion protein while maintaining its IgG binding activity through the Au-S bond. This enzyme-assisted one-pot methodology for protein-Au NPs conjugation offers one potent route for the facile fabrication of biomolecule-decorated metal NPs. © 2013 American Chemical Society.. |
| 219. |
Needle-free immunization using a solid-in-oil nanodispersion enhanced by skin-permeable oligoarginine peptide. |
| 220. |
Tailing DNA aptamers with a functional protein by two-step enzymatic reaction
An efficient, quantitative synthetic strategy for aptamer-enzyme conjugates was developed by using a two-step enzymatic reaction. Terminal deoxynucleotidyl transferase (TdT) was used to first incorporate a Z-Gln-Gly (QG) modified nucleotide which can act as a glutamine donor for a subsequent enzymatic reaction, to the 3'-OH of a DNA aptamer. Microbial transglutaminase (MTG) then catalyzed the cross-linking between the Z-QG modified aptamers and an enzyme tagged with an MTG-reactive lysine containing peptide. The use of a Z-QG modified dideoxynucleotide (Z-QG-ddUTP) or a deoxyuridine triphosphate (Z-QG-dUTP) in the TdT reaction enables the controlled introduction of a single or multiple MTG reactive residues. This leads to the preparation of enzyme-aptamer and (enzyme)n-aptamer conjugates with different detection limits of thrombin, a model analyte, in a sandwich enzyme-linked aptamer assay (ELAA). Since the combination of two enzymatic reactions yields high site-specificity and requires only short peptide substrates, the methodology should be useful for the labeling of DNA/RNA aptamers with proteins. © 2013 The Society for Biotechnology, Japan.. |
| 221. |
Hiromu Yoshiura, Miki Tamura, Makoto Aso, Noriho Kamiya, Masahiro Goto, Ionic liquid-in-oil microemulsions as potential carriers for the transdermal delivery of Methotrexate, JOURNAL OF CHEMICAL ENGINEERING OF JAPAN, 10.1252/jcej.13we009, 46, 11, 794-796, 2013.11, Ionic liquid-in-oil (IL/o) microemulsion (ME) techniques have been employed to increase the solubility of sparingly soluble drugs and enhance the extent of their topical and transdermal delivery. Methotrexate (MTX) has been used as a model drug in the current study. The size and size distributions of the ME droplets in the presence and in the absence of MTX were determined by dynamic light scattering analysis. The mean droplet diameter of the ME droplets containing 4 mg/mL of MTX was less than 20 nm, and smaller than the corresponding value observed in the absence of MTX. MTX was incorporated into IL droplets as well as other formulations and its permeation across Yucatan micropig (YMP) porcine skin evaluated. The use of IL/o ME has been shown to dramatically increase MTX administration. The results obtained in this study reveal the versatility of IL/o MEs as efficient drug delivery systems for sparingly soluble drugs.. |
| 222. |
Ionic liquid-in-oil microemulsions as potential carriers for the transdermal delivery of Methotrexate
Ionic liquid-in-oil (IL/o) microemulsion (ME) techniques have been employed to increase the solubility of sparingly soluble drugs and enhance the extent of their topical and transdermal delivery. Methotrexate (MTX) has been used as a model drug in the current study. The size and size distributions of the ME droplets in the presence and in the absence of MTX were determined by dynamic light scattering analysis. The mean droplet diameter of the ME droplets containing 4 mg/mL of MTX was less than 20 nm, and smaller than the corresponding value observed in the absence of MTX. MTX was incorporated into IL droplets as well as other formulations and its permeation across Yucatan micropig (YMP) porcine skin evaluated. The use of IL/o ME has been shown to dramatically increase MTX administration. The results obtained in this study reveal the versatility of IL/o MEs as efficient drug delivery systems for sparingly soluble drugs. © 2013 The Society of Chemical Engineers, Japan.. |
| 223. |
Aligning an endoglucanase Cel5A from Thermobifida fusca on a DNA scaffold: potent design of an artificial cellulosome. |
| 224. |
Biosorption of rare earth elements by Escherichia coli
Biosorption, which takes place by passive metal-sequestering, has recently attracted attention as an alternative to conventional separation processes such as precipitation and solvent extraction. In this work, the adsorption behavior of rare earth elements (REEs) onto an Escherichia coli (E. coli) bacterial adsorbent was examined. The adsorption of REEs increased with increasing pH of the feed solution. Also, E. coli showed a higher selectivity for heavier REEs, especially for scandium. To elucidate the adsorption mechanism, the functional groups on the E. coli were chemically modified. By analyzing the adsorption behavior and FT-IR spectra of these E. coli samples before and after adsorption, it was confirmed that REEs are adsorbed at phosphate and carboxyl groups on the surface of E. coli. It is concluded that E. coli is a potential novel adsorbent for REEs. © 2013 The Society of Chemical Engineers, Japan.. |
| 225. |
Tatsuaya Okuda, Tahara, Yoshiro, 神谷 典穂, Masahiro Goto, Satoru Kidoaki, S/O-nanodispersion electrospun fiber mesh effective for sustained release of healthy plasmid DNA with the structural and functional integrity, JOURNAL OF BIOMATERIALS SCIENCE-POLYMER EDITION, 10.1080/09205063.2012.755600, 24, 10, 1277-1290, 2013.07. |
| 226. |
Moriyama, Kousuke, Minamihata, Kosuke, Rie Wakabayashi, Masahiro Goto, 神谷 典穂, Enzymatic preparation of streptavidin-immobilized hydrogel using a phenolated linear poly(ethylene glycol), BIOCHEMICAL ENGINEERING JOURNAL, 10.1016/j.bej.2013.04.007, 76, 37-42, 2013.07. |
| 227. |
Yutaro Mori, Shiori Ozasa, Momoko Kitaoka, Shuhei Noda, Tsutomu Tanaka, Hirofumi Ichinose, Noriho Kamiya, Aligning an endoglucanase Cel5A from Thermobifida fusca on a DNA scaffold
Potent design of an artificial cellulosome, Chemical Communications, 10.1039/c3cc42614a, 49, 62, 6971-6973, 2013.07, A novel multi-cellulase conjugate assembled on a double-stranded DNA scaffold, a DNA–(endoglucanase)n conjugate, exhibited unique hydrolytic activity toward crystalline cellulose (Avicel) depending on the cellulase/DNA ratio on the DNA-based artificial cellulosome.. |
| 228. |
Enzymatic preparation of streptavidin-immobilized hydrogel using a phenolated linear poly(ethylene glycol)
Hybrid gels constructed from proteins and polymers have attracted a wide range of attention in the field of biomedicine and bioengineering. We report herein the enzymatic preparation of polymer-protein hybrid hydrogels composed of terminally bis-functionalized linear poly(ethylene glycol) (PEG) and streptavidin (SA). PEG was conjugated with tyramine to introduce terminal phenolic hydroxyl (Ph-OH) groups. A peptide tag containing a tyrosine residue (G4Y-tag) was genetically introduced at the C-terminus of SA. The Ph-OH-modified PEG and G4Y-tagged SA (SA-G4Y) were treated by horseradish peroxidase (HRP) in the presence of hydrogen peroxide (H2O2) to yield (PEG-Ph-OH)-(SA-G4Y) hybrid gels. Biotinylated enhanced green fluorescent protein (biotin-EGFP) was selectively captured in the obtained hybrid gels, indicating that SA-G4Y retained its biological function. The amount of biotin-EGFP immobilized in the hybrid gels depended on the concentration of SA-G4Y. In addition, biotinylated bacterial alkaline phosphatase (biotin-BAP) was immobilized in the hybrid gel. The immobilized biotin-BAP exhibited more than 95% of the initial activity after 5 rounds of recycling. The results suggest the facile functionalization of the hybrid gel with a variety of biotinylated functional molecules. © 2013 Elsevier B.V.. |
| 229. |
Uju, Abe, Kojiro, Uemura, Nobuyuki, Oshima, Toyoji, Masahiro Goto, 神谷 典穂, Peracetic acid-ionic liquid pretreatment to enhance enzymatic saccharification of lignocellulosic biomass, BIORESOURCE TECHNOLOGY, 10.1016/j.biortech.2013.03.147, 138, 87-94, 2013.06. |
| 230. |
Yang, Fan, Fukiko Kubota, Baba, Yuzo, 神谷 典穂, Masahiro Goto, Selective extraction and recovery of rare earth metals from phosphor powders in waste fluorescent lamps using an ionic liquid system, JOURNAL OF HAZARDOUS MATERIALS, 10.1016/j.jhazmat.2013.03.026, 254, 79-88, 2013.06. |
| 231. |
Muraoka, Jin;, 神谷 典穂, Ito, Yuji, Preparation and evaluation of cellulose-dissolving magnetic ionic liquid, JOURNAL OF MOLECULAR LIQUIDS, 10.1016/j.molliq.2013.03.012, 182, 76-78, 2013.06. |
| 232. |
Uju, Kojiro Abe, Nobuyuki Uemura, Toyoji Oshima, Masahiro Goto, Noriho Kamiya, Peracetic acid-ionic liquid pretreatment to enhance enzymatic saccharification of lignocellulosic biomass, BIORESOURCE TECHNOLOGY, 10.1016/j.biortech.2013.03.147, 138, 87-94, 2013.06, To enhance enzymatic saccharification of pine biomass, the pretreatment reagents peracetic acid (PAA) and ionic liquid (IL) were validated in single reagent pretreatments or combination pretreatments with different sequences. In a 1 h saccharification, 5-25% cellulose conversion was obtained from the single pretreatment of PAA or IL In contrast, a marked enhancement in conversion rates was achieved by PAA-IL combination pretreatments (45-70%). The PAA followed by IL (PAA + IL) pretreatment sequence was the most effective for preparing an enzymatic digestible regenerated biomass with 250-fold higher glucose formation rates than untreated biomass and 2- to 12-fold higher than single pretreatments with PAA or IL alone. Structural analysis confirmed that this pretreatment resulted in biomass with highly porous structural fibers associated with the reduction of lignin content and acetyl groups. Using the PAA + IL sequence, biomass loading in the pretreatment step can be increased from 5% to 15% without significant decrease in cellulose conversion. (C) 2013 Elsevier Ltd. All rights reserved.. |
| 233. |
Jin Muraoka, Noriho Kamiya, Yuji Ito, Preparation and evaluation of cellulose-dissolving magnetic ionic liquid, Journal of Molecular Liquids, 10.1016/j.molliq.2013.03.012, 182, 76-78, 2013.06, Ionic liquids have attracted attention as potential pretreatment agents in cellulosic biomass processing. Here we report on a new magnetic ionic liquid that can dissolve crystalline cellulose and be collected by a magnet. © 2013 Elsevier B.V.. |
| 234. |
Selective extraction and recovery of rare earth metals from phosphor powders in waste fluorescent lamps using an ionic liquid system
The recycling of rare earth metals from phosphor powders in waste fluorescent lamps by solvent extraction using ionic liquids was studied. Acid leaching of rare earth metals from the waste phosphor powder was examined first. Yttrium (Y) and europium (Eu) dissolved readily in the acid solution; however, the leaching of other rare earth metals required substantial energy input. Ionization of target rare earth metals from the waste phosphor powders into the leach solution was critical for their successful recovery. As a high temperature was required for the complete leaching of all rare earth metals, ionic liquids, for which vapor pressure is negligible, were used as an alternative extracting phase to the conventional organic diluent. An extractant, N, N-dioctyldiglycol amic acid (DODGAA), which was recently developed, showed a high affinity for rare earth metal ions in liquid-liquid extraction although a conventional commercial phosphonic extractant did not. An effective recovery of the rare earth metals, Y, Eu, La and Ce, from the metal impurities, Fe, Al and Zn, was achieved from the acidic leach solution of phosphor powders using an ionic liquid containing DODGAA as novel extractant system. © 2013 Elsevier B.V.. |
| 235. |
Uju, Shoda, Yasuhiro, Masahiro Goto, Tokuhara, Wataru, Ishida, Nobuhiro, Ogino, Chiaki, 神谷 典穂, Low melting point pyridinium ionic liquid pretreatment for enhancing enzymatic saccharification of cellulosic biomass, BIORESOURCE TECHNOLOGY, 10.1016/j.biortech.2012.06.096, 135, 103-108, 2013.05. |
| 236. |
Uju, Aya Nakamoto, Yasuhiro Shoda, Masahiro Goto, Wataru Tokuhara, Yoshiyuki Noritake, Satoshi Katahira, Nobuhiro Ishida, Chiaki Ogino, Noriho Kamiya, Low melting point pyridinium ionic liquid pretreatment for enhancing enzymatic saccharification of cellulosic biomass, BIORESOURCE TECHNOLOGY, 10.1016/j.biortech.2012.06.096, 135, 103-108, 2013.05, The potential of 1-hexylpyridinium chloride ([Hpy][Cl]), to pretreat cellulosic feedstocks was investigated using microcrystalline cellulose (Avicel) and Bagasse at 80 degrees C or 100 degrees C. Short [Hpy][Cl] pretreatments, |
| 237. |
Application of E. coli to adsorption and recovery of rare earth elements. |
| 238. |
Preparation of Multiple Emulsions to Depress the Rlease of Drugs and Enhanced Permeation Effect in Transdermal Delivery. |
| 239. |
Hiroki Abe, Rie Wakabayashi, Hiroaki Yonemura, Sunao Yamada, Masahiro Goto, Noriho Kamiya, Split Spy0128 as a Potent Scaffold for Protein Cross-Linking and Immobilization, BIOCONJUGATE CHEMISTRY, 10.1021/bc300606b, 24, 2, 242-250, 2013.02. |
| 240. |
Yutaro Mori, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya, Protein supramolecular complex formation by site-specific avidin-biotin interactions, Organic and Biomolecular Chemistry, 10.1039/c2ob26625c, 11, 6, 914-922, 2013.02, The precise accumulation of protein functions on a nanoscale to fabricate advanced biomaterials has become possible by a bottom-up approach based on molecular self-assembly. The avidin-biotin interaction is widely employed in the design of functional protein self-assemblies. Herein we assessed how the spatial arrangement of the avidin-biotin interaction between protein building blocks affects the formation of a protein supramolecular complex (PSC). The enzymatic site-specific internal labeling of a symmetric protein scaffold, bacterial alkaline phosphatase (AP), with specifically designed biotinylation substrates revealed that the precise positioning of the biotinylation sites on AP and the linker flexibility of the substrate are critical factors for the growth of PSCs in the presence of streptavidin (SA). A potential diagnostic application of the PSCs comprised of AP and SA was demonstrated in an enzyme-linked immunosorbent assay.. |
| 241. |
Protein supramolecular complex formation by site-specific avidin-biotin interactions
The precise accumulation of protein functions on a nanoscale to fabricate advanced biomaterials has become possible by a bottom-up approach based on molecular self-assembly. The avidin-biotin interaction is widely employed in the design of functional protein self-assemblies. Herein we assessed how the spatial arrangement of the avidin-biotin interaction between protein building blocks affects the formation of a protein supramolecular complex (PSC). The enzymatic site-specific internal labeling of a symmetric protein scaffold, bacterial alkaline phosphatase (AP), with specifically designed biotinylation substrates revealed that the precise positioning of the biotinylation sites on AP and the linker flexibility of the substrate are critical factors for the growth of PSCs in the presence of streptavidin (SA). A potential diagnostic application of the PSCs comprised of AP and SA was demonstrated in an enzyme-linked immunosorbent assay. © 2013 The Royal Society of Chemistry.. |
| 242. |
Split Spy0128 as a potent scaffold for protein cross-linking and immobilization
Site-specific cross-linking techniques between proteins and additional functional groups have become increasingly important for expanding the utility of proteins in biochemistry and biotechnology. In order to explore powerful techniques for practical bioconjugation applications, we have validated a technique mediated by a unique property of Streptcoccus pyogenes pilin subunit Spy0128, an autocatalytic intramolecular isopeptide formation in Spy0128. Recently, it has been revealed that Spy0128 can be split into two fragments (split-Spy0128 (residues 18-299 of Spy0128) and isopeptag (residues 293-308 of Spy0128)) that were capable of forming an intermolecular covalent complex. We focused on this unique reconstitution property and first studied the bioconjugation of blue and green fluorescent proteins, enabling the direct monitoring of cross-linking reactions by Förster resonance energy transfer (FRET). A fluorescence lifetime study shows that spatial control of two proteins on the Spy0128 scaffold is possible when one protein is fused to the N-terminus of split-Spy0128 and another one is tethered at the N- or C-terminus of the isopeptag. Furthermore, we demonstrated site-specific protein immobilization mediated by the reconstitution of split-Spy0128 and isopeptag. In this case, a split-Spy0128 mutant with a free N-terminal Cys residue was first immobilized onto beads chemically modified with a maleimide group through a Michael addition process. Then, an isopeptagged protein was successfully immobilized onto the split-Spy0128-immobilized beads. These results suggest that Spy0128 is a potent proteinaceous scaffold available for bioconjugation both in solution and at a solid surface. © 2013 American Chemical Society.. |
| 243. |
Development of Ionic Liquid Pharmaceuticals for Transdermal Drug Delivery. |
| 244. |
Shinya Furukawa, Kazuhiro Hasegawa, Ichino Fuke, Kkoji Kittaka, Terumitsu Nakakoba, Noriho Kamiya, Masahiro Goto, Enzymatic synthesis of Z-aspartame in liquefied amino acid substrates, BIOCHEMICAL ENGINEERING JOURNAL, 10.1016/j.bej.2012.10.002, 70, 84-87, 2013.01. |
| 245. |
Shinya Furukawa, Kazuhiro Hasegawa, Ichiro Fuke, Koji Kittaka, Terumitsu Nakakoba, Masahiro Goto, Noriho Kamiya, Enzymatic synthesis of Z-aspartame in liquefied amino acid substrates, Biochemical Engineering Journal, 10.1016/j.bej.2012.10.002, 70, 84-87, 2013.01, In this paper, we designed and validated a new media for biotransformation aided by ionic liquefied amino acid substrates. Thermolysin-catalyzed synthesis of Z-aspartame (N-carbobenzoxy-l-aspartame) in which ionic liquids play dual roles as both substrate and reaction medium was conducted. If the protease can retain catalytic activity in this new liquefied amino acid substrate/media, the effective substrate concentration can be maximized, leading to theoretically high yields. In fact, we observed a 34-fold enhancement in the catalytic activity compared with that obtained in a previous report. The yield of Z-aspartame reached at approximately 660mM at 24h with 50mgmL-1 of the enzyme, thus submolar range productivity was achieved. The results suggest the successful construction of a high productivity reaction system for peptide synthesis. © 2012 Elsevier B.V.. |
| 246. |
Tatsuya Okuda, Yoshiro Tahara, Noriho Kamiya, Masahiro Goto, Satoru Kidoaki, S/O-nanodispersion electrospun nanofiber mesh effective for sustained release of healthy plasmid DNA with the structuraland functional integrity, Journal of Biomaterials Science, Polymer Edition, 10.1080/09205063.2012.755600, 24, 10, 1277-1290, 2013.01. |
| 247. |
Fan Yang, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, A comparative study of ionic liquids and a conventional organic solvent on the extraction of rare-earth ions with TOPO, Solvent Extraction Research and Development, 10.15261/serdj.20.225, 20, 225-232, 2013, A comparative study of ionic liquids (ILs) and a conventional organic solvent on the extraction of rare-earth ions from a nitrate-contained solution with TOPO has been investigated. A slope analysis was carried out to determine the extraction mechanism, and it was found that more TOPO molecules and the ionic constituents of the ILs have taken part in the extraction reaction in the ILs-based TOPO extraction systems. The numbers of TOPO molecules coordinated to the metal ion in the extracted complexes were 6 and 4 in the [C4mim] and [C8mim][Tf2N] systems, although 3 TOPO molecules were needed in the n-dodecane system. Higher extraction efficiency was obtained in the extraction system in which the higher coordinating number of TOPO was shown. The extraction ability of [C8mim][Tf2N]-based TOPO depends significantly on the ionic constituents of the ILs in the extracting phase, but depends only slightly on the anionic nitrate ion from the aqueous phase, and [C4mim][Tf2N]-based TOPO is not dependent on the nitrate ion.. |
| 248. |
Yukiho Hosomomi, Yuzo Baba, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Biosorption of rare earth elements by Escherichia coli, JOURNAL OF CHEMICAL ENGINEERING OF JAPAN, 10.1252/jcej.13we031, 46, 7, 450-454, 2013, Biosorption, which takes place by passive metal-sequestering, has recently attracted attention as an alternative to conventional separation processes such as precipitation and solvent extraction. In this work, the adsorption behavior of rare earth elements (REEs) onto an Escherichia coli (E. coli) bacterial adsorbent was examined. The adsorption of REEs increased with increasing pH of the feed solution. Also, E. coli showed a higher selectivity for heavier REEs, especially for scandium. To elucidate the adsorption mechanism, the functional groups on the E. coli were chemically modified. By analyzing the adsorption behavior and FT-IR spectra of these E. coli samples before and after adsorption, it was confirmed that REEs are adsorbed at phosphate and carboxyl groups on the surface of E. coli. It is concluded that E. coli is a potential novel adsorbent for REEs.. |
| 249. |
Fan Yang, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Extraction of rare-earth ions with an 8-hydroxyquinoline derivative in an ionic liquid, Solvent Extraction Research and Development, 10.15261/serdj.20.123, 20, 123-129, 2013, The extraction behavior of rare-earth ions with an 8-hydroxyquinoline derivative, HO8Q (5-octyloxymethyl-8-quinolinol), was studied using an ionic liquid, [C8mim][Tf2N] (1-octyl-3-methylimidazolium bis(trifluoromethylsulfonyl)imide), as the extracting phase. Compared with a conventional organic solvent, n-dodecane, the ionic liquid system showed a higher extraction ability for a heavy rare earth, Dy, and a better selectivity between Dy and Nd (separation factor βDy/Nd: 108). In the liquid-liquid extraction system, the color of the extracting phase changed from colorless to light yellow along with the extraction of rare-earth ions. Furthermore, the extraction efficiency was enhanced by the addition of TOPO (tri-n-octylphosphine oxide) as a co-extractant.. |
| 250. |
Yoshiro Tahara, Shota Honda, Noriho Kamiya, Masahiro Goto, Transdermal delivery of insulin using a solid-in-oil nanodispersion enhanced by arginine-rich peptides, MEDCHEMCOMM, 10.1039/c2md20059g, 3, 12, 1496-1499, 2012.12. |
| 251. |
Yoshiro Tahara, Shota Honda, Noriho Kamiya, Masahiro Goto, Transdermal delivery of insulin using a solid-in-oil nanodispersion enhanced by arginine-rich peptides, MEDCHEMCOMM, 10.1039/c2md20059g, 3, 12, 1496-1499, 2012.12, A solid-in-oil nanodispersion that utilizes an oil-based vehicle of proteins was developed as a unique protein-delivery system into the skin. We enhanced the permeability of insulin and accomplished transdermal delivery of insulin by the collaborative effects of a S/O nanodispersion and arginine-rich peptides.. |
| 252. |
Yoshiro Tahara, Noriho Kamiya, Masahiro Goto, Solid-in-oil dispersion
A novel core technology for drug delivery systems, International Journal of Pharmaceutics, 10.1016/j.ijpharm.2012.09.007, 438, 1-2, 249-257, 2012.11, Drug delivery systems using a solid-in-oil (S/O) dispersion as a core technology have advanced significantly over the past ten years. A novel, effective and practical preparation method for a S/O dispersion was originally established in 1997 as a tool for enzymatic catalysis in organic media. This oil-based dispersion containing proteins in non-aqueous media had great potential for applications to other research with one of the most successful being its adaptation as a drug delivery system. The history and features of novel processes for preparing S/O dispersions are presented in this article. In addition, recent research into the use of S/O dispersions for innovative oral and skin drug delivery systems is discussed.. |
| 253. |
Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Activation of Pyrococcus furiosus alkaline phosphatase by divalent metal ions, BIOTECHNOLOGY LETTERS, 10.1007/s10529-012-0998-0, 34, 11, 2055-2060, 2012.11, Treatment of a hyperthermophilic enzyme, alkaline phosphatase from Pyrococcus furiosus (PfuAP), with EDTA completely deactivated PfuAP, indicating that the presence of one or more divalent metal ions is essential for its catalytic activity. Subsequent addition of various divalent metal ions to the apoprotein recovered the enzymatic activity and, in particular, the addition of Co(II) resulted in an over 50-fold increase in activity compared with PfuAP before EDTA treatment. Intriguingly, PfuAP with Co(II) exhibited weaker stability toward heat treatment, suggesting that Co2+ destabilizes the tertiary structure of PfuAP at high temperature.. |
| 254. |
Yoshiro Tahara, Noriho Kamiya, Masahiro Goto, Solid-in-oil dispersion: A novel core technology for drug delivery systems, INTERNATIONAL JOURNAL OF PHARMACEUTICS, 10.1016/j.ijpharm.2012.09.007, 438, 1-2, 249-257, 2012.11, Drug delivery systems using a solid-in-oil (S/O) dispersion as a core technology have advanced significantly over the past ten years. A novel, effective and practical preparation method for a S/O dispersion was originally established in 1997 as a tool for enzymatic catalysis in organic media. This oil-based dispersion containing proteins in non-aqueous media had great potential for applications to other research with one of the most successful being its adaptation as a drug delivery system. The history and features of novel processes for preparing S/O dispersions are presented in this article. In addition, recent research into the use of S/O dispersions for innovative oral and skin drug delivery systems is discussed. (c) 2012 Elsevier B.V. All rights reserved.. |
| 255. |
Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Activation of Pyrococcus furiosus alkaline phosphatase by divalent metal ions, Biotechnology letters, 10.1007/s10529-012-0998-0, 34, 11, 2055-2060, 2012.10, Treatment of a hyperthermophilic enzyme, alkaline phosphatase from Pyrococcus furiosus (PfuAP), with EDTA completely deactivated PfuAP, indicating that the presence of one or more divalent metal ions is essential for its catalytic activity. Subsequent addition of various divalent metal ions to the apoprotein recovered the enzymatic activity and, in particular, the addition of Co(II) resulted in an over 50-fold increase in activity compared with PfuAP before EDTA treatment. Intriguingly, PfuAP with Co(II) exhibited weaker stability toward heat treatment, suggesting that Co2+ destabilizes the tertiary structure of PfuAP at high temperature.. |
| 256. |
Muhammad Moniruzzaman, Keishirou Ino, Noriho Kamiya, Masahiro Goto, Lipase incorporated ionic liquid polymers as active, stable and reusable biocatalysts, Organic and Biomolecular Chemistry, 10.1039/c2ob25529d, 10, 38, 7707-7713, 2012.10, The aim of this study was to develop ionic liquid (IL) polymer materials incorporating enzymes that can be used as active, stable and reusable biocatalysts. To this goal, Candida rugosa lipase has been microencapsulated in surfactant aggregates formed in an IL monomer or the solution of an IL monomer/IL and then incorporated into polymer frameworks through the free radical polymerization of an IL (1-vinyl-3-ethylimidazolium bis(trifluoromethyl- sulfonyl) amide) ([veim][Tf2N]). The activity, stability and reusability of such IL polymer materials containing lipase were evaluated using lipase-catalyzed hydrolysis of p-nitrophenyl butyrate (p-PNB) as a model reaction. Lipase encapsulated within ionic liquid polymer materials remained active and exhibited excellent stability in aqueous solutions. More importantly, these biopolymer materials retained most of their activity after five reaction cycles, in which biopolymers were recovered from the reaction mixture simply by centrifugation. This study promulgates a direction toward the design of IL-an interesting class of tunable and designable solvents-based polymer materials containing biomolecules via a combination of polymer and supramolecular chemistry for diverse applications.. |
| 257. |
Fan Yang, Noriho Kamiya, Masahiro Goto, Transdermal delivery of the anti-rheumatic agent methotrexate using a solid-in-oil nanocarrier, EUROPEAN JOURNAL OF PHARMACEUTICS AND BIOPHARMACEUTICS, 10.1016/j.ejpb.2012.05.016, 82, 1, 158-163, 2012.09. |
| 258. |
Fan Yang, Noriho Kamiya, Masahiro Goto, Transdermal delivery of the anti-rheumatic agent methotrexate using a solid-in-oil nanocarrier, EUROPEAN JOURNAL OF PHARMACEUTICS AND BIOPHARMACEUTICS, 10.1016/j.ejpb.2012.05.016, 82, 1, 158-163, 2012.09, Transdermal delivery of methotrexate (MIX) was investigated by using the solid-in-oil (S/O) technique. Because MTX was coated with nonionic surfactant molecules, the resulting complex was easy to dissolve in various organic solvents and provided a transparent solution in isopropyl myristate (IPM). The stability of MIX-surfactant complexes are enhanced by the addition of a basic amino acid such as L-Arginine (L-Arg) or L-Lysine (L-Lys). The average size of the dispersed complex of MIX and amino acid was reduced to below 100 nm and gave a uniform distribution. A transdermal delivery experiment was conducted using the S/O nanocarrier, and the permeation behavior of MIX through Yucatan micropig (YMP) skin was evaluated with a Franz diffusion cell. The permeation efficiency for the S/O nanocarrier (not urea addition) was two- to threefold increased compared to that of the control aqueous solution because the oil-based nanocarrier is effective for penetrating the stratum corneum. Furthermore, addition of urea has dramatically improved the release property of MIX from the S/O nanocarrier, and the S/O nanocarrier containing urea showed an optimal permeation efficiency of approximately 8.8-fold increased compared to that of the control aqueous solution after 24 h (p |
| 259. |
Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Control of a Tyrosyl Radical Mediated Protein Cross-Linking Reaction by Electrostatic Interaction, BIOCONJUGATE CHEMISTRY, 10.1021/bc300137s, 23, 8, 1600-1609, 2012.08. |
| 260. |
Muhammad Moniruzzaman, Keishirou Ino, Noriho Kamiya, Masahiro Goto, Lipase incorporated ionic liquid polymers as active, stable and reusable biocatalysts, Org. Biomol. Chem., 14, 7707-7713, 2012.08. |
| 261. |
Yoshiro Tahara, Takeshi Kaneko, Riki Toita, Chiharu Yoshiyama, Takuya Kitaoka, Takuro Niidome, Yoshiki Katayama, Noriho Kamiya, Masahiro Goto, A novel double-coating carrier produced by solid-in-oil and solid-in-water nanodispersion technology for delivery of genes and proteins into cells, JOURNAL OF CONTROLLED RELEASE, 10.1016/j.jconrel.2012.05.001, 161, 3, 713-721, 2012.08. |
| 262. |
Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Activation of Pyrococcus furiosus alkaline phosphatase by divalent metal ions, Biotechnol. Lett., 34, 2055-2060, 2012.08. |
| 263. |
Yoshiro Tahara, Takeshi Kaneko, Riki Toita, Chiharu Yoshiyama, Takuya Kitaoka, Takuro Niidome, Yoshiki Katayama, Noriho Kamiya, Masahiro Goto, A novel double-coating carrier produced by solid-in-oil and solid-in-water nanodispersion technology for delivery of genes and proteins into cells, JOURNAL OF CONTROLLED RELEASE, 10.1016/j.jconrel.2012.05.001, 161, 3, 713-721, 2012.08, A novel intracellular delivery method both for genes and proteins is one of the most coveted systems in the drug delivery field. In the present study, we developed a double-coating carrier loaded with gene and protein produced by solid-in-oil and solid-in-water nanodispersion technology. The double-coating carriers did not require electrostatic interactions during the preparation so were able to encapsulate plasmid DNA, ovalbumin (pI 4.5), horseradish peroxidase (pI 7.2), and cytochrome-c (pI 10.5) in a consistent manner. The carriers had practical encapsulation efficiencies and release profiles for genes and proteins. Furthermore, effective gene expression and cellular uptakes of both anionic and cationic proteins were achieved by modification of carriers with functional molecules. These findings indicate that the double-coating carrier has high potential for cellular delivery of various drugs and is a novel, superior method for both gene and protein delivery into cells. (c) 2012 Elsevier B.V. All rights reserved.. |
| 264. |
Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Control of a Tyrosyl Radical Mediated Protein Cross-Linking Reaction by Electrostatic Interaction, BIOCONJUGATE CHEMISTRY, 10.1021/bc300137s, 23, 8, 1600-1609, 2012.08, Herein, we demonstrate the control of protein heteroconjugation via a tyrosyl coupling reaction by using electrostatic interaction. Aspartic acid and arginine were introduced. to a tyrosine containing peptide tag (Y-tag) to provide electrostatic charge. Designed negatively or positively charged Y-tags were tethered to the C-terminus of Escherichia coli alkaline phosphatase (BAP) and streptavidin (SA), and these model proteins were subjected to horseradish peroxidase (HRP) treatment The negatively charged Y-tags showed low reactivity due to repulsive interactions between the Y-tags with the negatively charged BAP and SA. In contrast, the positively charged Y-tags showed high reactivity, indicating that the electrostatic interaction between Y-tags and proteins significantly affects the tyrosyl radical mediated protein cross linking From the heteroconjugation reaction of BAP and SA, the SA with the positively charged Y-tags exhibited favorable cross linking toward negatively charged BAP, and the BAP SA conjugates prepared from BAP with GY-tag (GGGGY) and SA with RYR-tag (RRYRR) had the best performance on a biotin coated microplate. Encompassing the reactive tyrosine residue with arginine residues reduced the reactivity against HRP, enabling the modulation of cross linking reaction rates with BAP-GY. Thus, by introducing a proper electrostatic interaction to Y-tags, it is possible to kinetically control the heteroconjugation behavior of proteins, thereby maximizing the functions of protein heteroconjugates.. |
| 265. |
Noriho Kamiya, Momoko Kitaoka, Kounosuke Hayashi, Yoshiyuki HIraishi, Koji Nakano, Katsuyuki Miyawaki, Sumihare Noji, Masahiro Goto, Transglutaminase-Mediated in Situ Hybridization (TransISH) System: A New Methodology for Simplified mRNA Detection, ANALYTICAL CHEMISTRY, 10.1021/ac2034198, 84, 14, 5885-5891, 2012.07, In situハイブリダイゼーション法は遺伝子機能解明のための重要な基礎的技術となっているが、免疫染色技術を利用する一般的な方法では、煩雑で時間のかかる操作が必要とされる。本研究では、簡易な操作でターゲット核酸を検出するための新たなRNA―酵素複合体の調製方法を開発し、それを用いた組織切片上での高感度核酸検出を達成した。. |
| 266. |
Momoko Kitaoka, Masayuki Mitsumori, Kounosuke Hayashi, Yoshiyuki Hiraishi, Hisao Yoshinaga, Koji Nakano, Katsuyuki Miyawaki, Sumihare Noji, Masahiro Goto, Noriho Kamiya, Transglutaminase-Mediated in Situ Hybridization (TransISH) System: A New Methodology for Simplified mRNA Detection, ANALYTICAL CHEMISTRY, 10.1021/ac2034198, 84, 14, 5885-5891, 2012.07, Detection and localization of specific DNA or RNA sequences in cells and tissues are of great importance for biological research, diagnosis, and environmental monitoring. However, the most common procedure for in situ hybridization employs laborious immunostaining techniques. In the present study, we report proof-of-concept for a new RNA enzyme conjugated probe for the detection of mRNA on tissue sections with a simple procedure. An RNA probe modified with a specific dipeptide substrate of transglutaminase was prepared. Alkaline phosphatase was then covalently and site specifically combined to the dipeptide-labeled RNA using microbial transglutaminase. The new RNA probe labeled with alkaline phosphatase-was validated by in situ hybridization (ISH) and proved to be a sensitive and sequence specific probe for mRNA detection in tissues.. The new transglutaminase-mediated ISH (TransISH) strategy is free from antigen-antibody reaction, leads to one-step signal amplification after hybridization, and thus will be widely applicable for highly sensitive nucleic acid detection.. |
| 267. |
Kojiro Shimojo, Teppei Niide, Tomitsugu Taguchi, Hirochika Naganawa, Noriho Kamiya, Masahiro Goto, Facile, rapid and efficient biofabrication of gold nanoparticles decorated with functional proteins, Analyst, 10.1039/c2an35172b, 137, 10, 2300-2303, 2012.05, We report a one-pot biological approach to fabricate gold nanoparticle (AuNP)-ZZ domain conjugates using peptide-functionalized proteins that can simultaneously direct both biomineralization and surface modification of AuNPs. In addition, immuno-AuNPs are readily prepared through the specific binding of antibodies to the ZZ domain on the AuNPs.. |
| 268. |
Josui Shimada, Tatsuo Maruyama, Momoko Kitaoka, Hisao Yoshinaga, Koji Nakano, Noriho Kamiya, Masahiro Goto, Programmable protein-protein conjugation via DNA-based self-assembly, Chemical Communications, 10.1039/c2cc30618b, 48, 50, 6226-6228, 2012.05, Protein molecules were precisely arrayed on a designable DNA scaffold close to each other using a DNA aptamer. By adding a chemical cross-linker, the neighboring protein molecules were effectively and covalently cross-linked to each other without losing their activities.. |
| 269. |
J.Shimada, T. Maruyama, M. Kitaoka, N. Kamiya, M. Goto, Microplate assay for aptamer-based thrombin detection using a DNA-enzyme conjugate based on His-tag chemistry, Anal. Biochem., 421, 541-546, 2012.02. |
| 270. |
Eiichi Toorisaka, Kikumi Watanabe, Hiroshige Ono, Makoto Hirata, Noriho Kamiya, Masahiro Goto, Intestinal patches with an immobilized solid-in-oil formulation for oral protein delivery, Acta Biomaterialia, 10.1016/j.actbio.2011.09.023, 8, 2, 653-658, 2012.02, Oral administration of biomolecular drugs such as peptides, proteins, and DNA is an attractive delivery method because of the safety and convenience of delivery in contrast to injection administration. However, oral delivery of biomolecules has several potential barriers such as enzymatic degradation in the gastrointestinal tract and low permeability across an intestinal membrane. In this study, we proposed an intestinal patch system that included surfactant-coated insulin for oral delivery. The intestinal patches, which have mucoadhesive and drug-impermeable layers, induced sustained unidirectional insulin release toward intestinal mucosa and inhibition of insulin leakage from the patches. Moreover, the surfactant-coated insulin, which has high compatibility with cell membranes, enhanced insulin transport across the intestinal membrane. This study demonstrates that the intestinal patches might improve protein permeability in the intestinal mucosa, thereby offering an innovative therapeutic strategy.. |
| 271. |
Josui Shimada, Tatsuo Maruyama, Momoko Kitaoka, Noriho Kamiya, Masahiro Goto, Microplate assay for aptamer-based thrombin detection using a DNA-enzyme conjugate based on histidine-tag chemistry, Analytical Biochemistry, 10.1016/j.ab.2011.11.028, 421, 2, 541-546, 2012.02, We report a method to prepare a DNA-enzyme conjugate using histidine-tag (His-tag) chemistry. A DNA oligonucleotide was modified with nitrilotriacetate (NTA), whose Kd was approximately 10-6 (M-1) toward a His-tag present on a recombinant protein via the complexation of Ni2+. His-tagged alkaline phosphatase (His-AP) was used as the model enzyme. Enzyme immobilization on the microplate revealed the conjugation of His-AP and the NTA-modified DNA via an Ni2+ complex. SPR measurements also proved the conjugation of His-AP with the NTA-modified DNA via an Ni 2+ complex. The DNA-enzyme conjugate was then used for the detection of thrombin using a DNA aptamer. The DNA-AP conjugate successfully amplified the binding signal between the DNA aptamer and the thrombin, and the signal was measured as the fluorescent intensity derived from the AP-catalyzed reaction. The detection limit was 11 nM. Finally, we studied the effect of the release of the immobilized His-AP from the microplate on the AP activity, because the present strategy used a cleavable linker for the conjugation and the enzyme immobilization. The DNase-catalyzed release of the immobilized His-AP resulted in a 1.7-fold higher AP activity than observed when the His-AP was surface-immobilized.. |
| 272. |
Eiichi Toorisaka, Kikumi Watanabe, Hiroshige Ono, Makoto Hirata, Noriho Kamiya, Masahiro Goto, Intestinal patches with an immobilized solid-in-oil formulation for oral protein delivery, ACTA BIOMATERIALIA, 10.1016/j.actbio.2011.09.023, 8, 2, 653-658, 2012.02, Oral administration of biomolecular drugs such as peptides, proteins, and DNA is an attractive delivery method because of the safety and convenience of delivery in contrast to injection administration. However, oral delivery of biomolecules has several potential barriers such as enzymatic degradation in the gastrointestinal tract and low permeability across an intestinal membrane. In this study, we proposed an intestinal patch system that included surfactant-coated insulin for oral delivery. The intestinal patches, which have mucoadhesive and drug-impermeable layers, induced sustained unidirectional insulin release toward intestinal mucosa and inhibition of insulin leakage from the patches. Moreover, the surfactant-coated insulin, which has high compatibility with cell membranes, enhanced insulin transport across the intestinal membrane. This study demonstrates that the intestinal patches might improve protein permeability in the intestinal mucosa, thereby offering an innovative therapeutic strategy. (C) 2011 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.. |
| 273. |
Josui Shimada, Tatsuo Maruyama, Momoko Kitaoka, Noriho Kamiya, Masahiro Goto, Microplate assay for aptamer-based thrombin detection using a DNA-enzyme conjugate based on histidine-tag chemistry, ANALYTICAL BIOCHEMISTRY, 10.1016/j.ab.2011.11.028, 421, 2, 541-546, 2012.02, We report a method to prepare a DNA-enzyme conjugate using histidine-tag (His-tag) chemistry. A DNA oligonucleotide was modified with nitrilotriacetate (NTA), whose K-d was approximately 10(-6) (M-1) toward a His-tag present on a recombinant protein via the complexation of Ni2+. His-tagged alkaline phosphatase (His-AP) was used as the model enzyme. Enzyme immobilization on the microplate revealed the conjugation of His-AP and the NTA-modified DNA via an Ni2+ complex. SPR measurements also proved the conjugation of His-AP with the NTA-modified DNA via an Ni2+. complex. The DNA-enzyme conjugate was then used for the detection of thrombin using a DNA aptamer. The DNA-AP conjugate successfully amplified the binding signal between the DNA aptamer and the thrombin, and the signal was measured as the fluorescent intensity derived from the AP-catalyzed reaction. The detection limit was 11 nM. Finally, we studied the effect of the release of the immobilized His-AP from the microplate on the AP activity, because the present strategy used a cleavable linker for the conjugation and the enzyme immobilization. The DNase-catalyzed release of the immobilized His-AP resulted in a 1.7-fold higher AP activity than observed when the His-AP was surface-immobilized. (C) 2011 Elsevier Inc. All rights reserved.. |
| 274. |
チロシルラジカルの電荷指向型カップリング反応によるタンパク質架橋. |
| 275. |
チロシンタグ導入タンパク質/フェノール性水酸基含有ヒドロゲル融合基材の開発. |
| 276. |
Uju, Yasuhiro Shoda, Aya Nakamoto, Masahiro Goto, Wataru Tokuhara, Yoshiyuki Noritake, Satoshi Katahira, Nobuhiro Ishida, Kazunori Nakashima, Chiaki Ogino, Noriho Kamiya, Short time ionic liquids pretreatment on lignocellulosic biomass to enhance enzymatic saccharification, Bioresource Technology, 10.1016/j.biortech.2011.10.003, 103, 1, 446-452, 2012.01, The potential of 1-buthyl-3-methylpyridinium chloride, [Bmpy][Cl], as a pretreatment solvent for lignocellulosic biomasses, Bagasse and Eucalyptus, was investigated. The yields of regenerated biomasses ranged between 35% and 96%, and varied according to the pretreatment time, type of ionic liquid (IL) and biomass. The pretreatment of the biomass with [Bmpy][Cl] resulted in up to 8-fold increase in the cellulose conversion when compared with the untreated biomass. For a short pretreatment period (i.e., 10. min), [Bmpy][Cl] showed better performance than 1-ethyl-3-methylimidazolium acetate ([Emim][OAc]) with respect to the initial enzymatic saccharification rates. The increase in the reaction rates with [Emim][OAc] treatment was because of a reduction in the cellulose crystallinity. In contrast, a decrease in the crystallinity index was not clearly observed for the biomass pretreated with [Bmpy][Cl], and the enhancement of the enzymatic saccharification rates using this IL is presumably due to a reduction in the degree of polymerization of cellulose in the biomass.. |
| 277. |
Uju, Yasuhiro Shoda, Aya Nakamoto, Masahiro Goto, Wataru Tokuhara, Yoshiyuki Noritake, Satoshi Katahira, Nobuhiro Ishida, Kazunori Nakashima, Chiaki Ogino, Noriho Kamiya, Short time ionic liquids pretreatment on lignocellulosic biomass to enhance enzymatic saccharification, BIORESOURCE TECHNOLOGY, 10.1016/j.biortech.2011.10.003, 103, 1, 446-452, 2012.01, The potential of 1-buthyl-3-methylpyridinium chloride, [Bmpy][Cl], as a pretreatment solvent for lignocellulosic biomasses, Bagasse and Eucalyptus, was investigated. The yields of regenerated biomasses ranged between 35% and 96%, and varied according to the pretreatment time, type of ionic liquid (IL) and biomass. The pretreatment of the biomass with [Bmpy][Cl] resulted in up to 8-fold increase in the cellulose conversion when compared with the untreated biomass. For a short pretreatment period (i.e., 10 min), [Bmpy][Cl] showed better performance than 1-ethyl-3-methylimidazolium acetate ([Emim][OAc]) with respect to the initial enzymatic saccharification rates. The increase in the reaction rates with [Emim][OAc] treatment was because of a reduction in the cellulose crystallinity. In contrast, a decrease in the crystallinity index was not clearly observed for the biomass pretreated with [Bmpy][Cl], and the enhancement of the enzymatic saccharification rates using this IL is presumably due to a reduction in the degree of polymerization of cellulose in the biomass. (C) 2011 Published by Elsevier Ltd.. |
| 278. |
Fan Yang, Yuzo Baba, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Extraction and separation of rare earth metal ions with DODGAA in ionic liquids, Solvent Extraction Research and Development, 10.15261/serdj.19.69, 19, 69-76, 2012, In order to apply ionic liquids (ILs) to the separation of rare earth metals, liquid-liquid extraction of a series of rare earth elements into the ILs was investigated mainly using 1-octyl-3-methylimidazolium bis(trifluoromethyl sulfonyl) imide, [C8mim][Tf2N], with N,N-dioctyldiglycol amic acid (DODGAA) as the extractant. A higher selectivity was exhibited for the heavier rare earth elements as in the n-dodecane system, and the order of the extractability of Y(III) in the IL system, situated in the heavy rare earth metals, was between Tb and Dy. This position is a little different from that for the common solvent extraction system employing organophosphorus extractants such as bis (2-ethylhexyl)phosphoric acid (D2EHPA) and 2-ethylhexylphosphonic acid mono-2-ethylhexyl ester (PC-88A). Quantitative extraction was achieved for all rare earth metals with DODGAA from sulfuric acid solutions from pH 1.5 to 4. It was confirmed that rare earth metal ions were extracted with 3 DODGAA molecules in the ILs.. |
| 279. |
H. Abe, M. Goto, N. Kamiya, Protein lipidation catalyzed by microbial transglutaminase, Chem. Eur. J., 17, 14004-14008, 2011.12. |
| 280. |
K. Minamihata, M. Goto, N. Kamiya, Protein heteroconjugation by the peroxidase-catalyzed tyrosine coupling reaction, Bioconjugate Chem., 22, 2332-2338, 2011.12. |
| 281. |
Hiroki Abe, Masahiro Goto, Noriho Kamiya, Protein lipidation catalyzed by microbial transglutaminase, Chemistry - A European Journal, 10.1002/chem.201102121, 17, 50, 14004-14008, 2011.12, Fattening up: Artificially constructed lipid-protein conjugates can be used to control protein activity and localization at either biological or artificial membrane interfaces. Here, we report a facile technique for the lipid modification of proteins catalyzed by microbial transglutaminase (MTG). MTG-mediated protein lipidation proceeded efficiently without perturbing protein functionality (see figure).. |
| 282. |
Hiroki Abe, Masahiro Goto, Noriho Kamiya, Protein Lipidation Catalyzed by Microbial Transglutaminase, CHEMISTRY-A EUROPEAN JOURNAL, 10.1002/chem.201102121, 17, 50, 14004-14008, 2011.12, Fattening up: Artificially constructed lipid-protein conjugates can be used to control protein activity and localization at either biological or artificial membrane interfaces. Here, we report a facile technique for the lipid modification of proteins catalyzed by microbial transglutaminase (MTG). MTG-mediated protein lipidation proceeded efficiently without perturbing protein functionality (see figure). Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.. |
| 283. |
Hongyu Piao, Noriho Kamiya, Fude Cui, Masahiro Goto, Preparation of a solid-in-oil nanosuspension containing l-ascorbic acid as a novel long-term stable topical formulation, International Journal of Pharmaceutics, 10.1016/j.ijpharm.2011.08.025, 420, 1, 156-160, 2011.11, l-Ascorbic acid (AA, vitamin C) easily decomposes into inactive compounds in aqueous solutions and this has limited its topical use. This work reports the preparation of a solid-in-oil nanosuspension (SONS) containing AA and validation of its basic storage stability. Although AA itself is water-soluble, it can readily be nanosuspended in squalane via complex formation involving a combination of sucrose erucate (i.e. lipophilic surfactant) and sucrose monolaureate (i.e. hydrophilic surfactant) to yield SONS with a very low moisture content ( |
| 284. |
Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Protein heteroconjugation by the peroxidase-catalyzed tyrosine coupling reaction, Bioconjugate Chemistry, 10.1021/bc200420v, 22, 11, 2332-2338, 2011.11, Combining different proteins can integrate the functions of each protein to produce novel protein conjugates with wider ranges of applications. We have previously introduced a peptide containing tyrosine residues (Y-tag) at the C-terminus of Escherichia coli alkaline phosphatase (BAP). The tyrosine residues in the Y-tag were efficiently recognized by horseradish peroxidase (HRP) and were site-specifically cross-linked with each other to yield BAP homoconjugates. In this study, the HRP-catalyzed tyrosine coupling reaction was used for protein heteroconjugation. Streptavidin (SA) was selected as the conjugation partner for BAP. The Y-tag (GGGGY) was genetically introduced to the C-terminus of SA. Prior to heteroconjugation, the reactivity of the Y-tagged SA was examined. The Y-tagged SA cross-linked to form an SA homoconjugate upon HRP treatment, whereas wild-type SA remained essentially intact. In the heteroconjugation reaction of BAP and SA, the Y-tagged BAP and SA were efficiently cross-linked with each other upon HRP treatment. The functions of the BAP-SA conjugates were evaluated by measuring the BAP enzymatic activity on a biotin-coated plate. The BAP-SA conjugate tethered to the plate showed BAP enzymatic activity, indicating that both BAP and SA retained their functions following heteroconjugation. The BAP-SA conjugate prepared from both Y-tagged BAP and SA showed the highest enzymatic activity on the biotin-coated plates. This result illustrates the advantage of the protein conjugation reaction in which multiple numbers of proteins can be conjugated at the same time.. |
| 285. |
Yuzo Baba, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Recent advances in extraction and separation of rare-earth metals using ionic liquids, JOURNAL OF CHEMICAL ENGINEERING OF JAPAN, 10.1252/jcej.10we279, 44, 10 SPEC. ISSUE, 679-685, 2011.11, In the present review article, we summarize recent advances in extraction and separation of rare-earth metals using ionic liquids. Rare-earth metals have unique electronic and magnetic properties. In Japan, increasing amounts of rare-earth metals are being used every year in industries employing cutting-edge technology. Therefore, maintaining a stable supply of rare-earth metals is important. In recent times, extensive research has been carried out on the use of ionic liquids for the extraction of rare-earth metals; this is because the extraction ability and/or selectivity for rare-earth metals have been found to increase when using hydrophobic ionic liquids, which have potential applications in liquid-liquid extraction processes. In the octyl(phenyl)-N,N-diisobutyl carbamoylmethyl phosphine oxide (CMPO) extraction system using ionic liquids, the extraction ability was more than 103 times better than that when using n-dodecane; however, stripping of the extracted metal ions was difficult in the former case. In initial research on the extraction processes based on ionic liquids, industrial extractant PC-88A was employed as well as CMPO, but issues resulting from the low solubility of these extractants and their metal complexes remain to be addressed. To overcome this problem, a new extractant, N,N-dioctyldiglycol amic acid (DODGAA), which is highly soluble in the ionic liquids, has been synthesized. DODGAA shows better performance for the separation of rare-earth metals in ionic liquids than in n-dodecane. Furthermore, a liquid membrane system based on ionic liquids has been developed for the separation and recovery of rare-earth metals. The use of a liquid membrane system may help in reducing the amount of ionic liquids required for metal recovery.. |
| 286. |
Hongyu Piao, Noriho Kamiya, Fude Cui, Masahiro Goto, Preparation of a solid-in-oil nanosuspension containing L-ascorbic acid as a novel long-term stable topical formulation, INTERNATIONAL JOURNAL OF PHARMACEUTICS, 10.1016/j.ijpharm.2011.08.025, 420, 1, 156-160, 2011.11, L-Ascorbic acid (AA, vitamin C) easily decomposes into inactive compounds in aqueous solutions and this has limited its topical use. This work reports the preparation of a solid-in-oil nanosuspension (SONS) containing AA and validation of its basic storage stability. Although AA itself is water-soluble, it can readily be nanosuspended in squalane via complex formation involving a combination of sucrose erucate (i.e. lipophilic surfactant) and sucrose monolaureate (i.e. hydrophilic surfactant) to yield SONS with a very low moisture content ( |
| 287. |
Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Protein Heteroconjugation by the Peroxidase-Catalyzed Tyrosine Coupling Reaction, BIOCONJUGATE CHEMISTRY, 10.1021/bc200420v, 22, 11, 2332-2338, 2011.11, Combining different proteins can integrate the functions of each protein to produce novel protein conjugates with wider ranges of applications. We have previously introduced a peptide containing tyrosine residues (Y-tag) at the C-terminus of Escherichia coli alkaline phosphatase (BAP). The tyrosine residues in the Y-tag were efficiently recognized by horseradish peroxidase (HRP) and were site-specifically cross-linked with each other to yield BAP homoconjugates. In this study, the HRP-catalyzed tyrosine coupling reaction was used for protein heteroconjugation. Streptavidin (SA) was selected as the conjugation partner for BAP. The Y-tag (GGGGY) was genetically introduced to the C-terminus of SA. Prior to heteroconjugation, the reactivity of the Y-tagged SA was examined. The Y-tagged SA cross-linked to form an SA homoconjugate upon HRP treatment, whereas wild-type SA remained essentially intact. In the heteroconjugation reaction of BAP and SA, the Y-tagged BAP and SA were efficiently cross-linked with each other upon FIR? treatment. The functions of the BAP-SA conjugates were evaluated by measuring the BAP enzymatic activity on a biotin-coated plate. The BAP-SA conjugate tethered to the plate showed BAP enzymatic activity, indicating that both BAP and SA retained their functions following heteroconjugation. The BAP-SA conjugate prepared from both Y-tagged BAP and SA showed the highest enzymatic activity on the biotin-coated plates. This result illustrates the advantage of the protein conjugation reaction in which multiple numbers of proteins can be conjugated at the same time.. |
| 288. |
H. Piao, N. Kamiya, F. Cui, M. Goto, Preparation of a solid-in-oil nanosuspension containing l-ascorbic acid as a novel long-term stable topical formulation, Int. J. Pharm., 420, 156-160, 2011.10. |
| 289. |
J.Shimada, T. Maruyama, M. Kitaoka, N. Kamiya, M. Goto, DNA-enzyme conjugate with a weak inhibitor that can specifically detect thrombin in a homogeneous medium, Anal. Biochem., 414, 103-108, 2011.09. |
| 290. |
Yutaro Mori, Kosuke Minamihata, Hiroki Abe, Masahiro Goto, Noriho Kamiya, Protein assemblies by site-specific avidin-biotin interactions, Organic and Biomolecular Chemistry, 10.1039/c1ob05428g, 9, 16, 5641-5644, 2011.08, Exploiting self-assembly systems with biological building blocks is of significant interest in the fabrication of advanced biomaterials. We assessed the potential use of site-specific ligand labeling of protein building blocks in designing functional protein self-assemblies by combining site-specifically biotinylated bacterial alkaline phosphatase (as a bidentate or tetradentate ligand unit) and streptavidin (as a tetrameric receptor).. |
| 291. |
ストレプトアビジンー酵素ハイブリッドの新規調製法の開発. |
| 292. |
Teppei Niide, Masahiro Goto, Noriho Kamiya, Biocatalytic synthesis of gold nanoparticles with cofactor regeneration in recombinant Escherichia coli cells, Chemical Communications, 10.1039/c1cc12302e, 47, 26, 7350-7352, 2011.07, Here we report the enzymatic synthesis of gold nanoparticles (Au NPs) by an engineered Escherichia coli harboring an NADH cofactor regeneration system coupled with glycerol dehydrogenase, which can be triggered by the addition of exogenous glycerol.. |
| 293. |
Josui Shimada, Tatsuo Maruyama, Momoko Kitaoka, Noriho Kamiya, Masahiro Goto, DNA-enzyme conjugate with a weak inhibitor that can specifically detect thrombin in a homogeneous medium, Analytical Biochemistry, 10.1016/j.ab.2011.02.035, 414, 1, 103-108, 2011.07, We present the DNA-assisted control of enzymatic activity for the detection of a target protein using a new type of DNA-enzyme conjugate. The conjugate is composed of an enzyme inhibitor to regulate enzyme activity and a DNA aptamer to be responsive toward the analyte protein. Glutathione S-transferase (GST) and thrombin were selected as a model enzyme and an analyte protein. A hexahistidine tag was genetically attached to the C terminus of the GST, and the 5′ end of an oligonucleotide was conjugated with nitrilotriacetic acid (NTA) for the site-specific conjugation of the DNA with the GST based on a Ni2+ complex interaction. We found that fluorescein acted as a weak inhibitor of GST and succeeded in the regulation of GST activity by increasing the local concentration of the weak inhibitor by the hybridization of a 3′-end fluorescein-modified DNA. The catalytic activity of the DNA aptamer-enzyme conjugate showed a dose-dependent response to thrombin, indicating that the GST activity was clearly recovered by the binding of the DNA aptamer to thrombin. The current system enables the sensitive and specific detection of thrombin simply by measuring the enzymatic activity in a homogeneous medium.. |
| 294. |
Yutaro Mori, Masahiro Goto, Noriho Kamiya, Transglutaminase-mediated internal protein labeling with a designed peptide loop, Biochemical and Biophysical Research Communications, 10.1016/j.bbrc.2011.06.073, 410, 4, 829-833, 2011.07, Post-translational internal protein labeling was explored through the insertion of a 13-mer peptidyl loop specifically recognized by microbial transglutaminase (MTG). The peptidyl loop included one lysine residue (abbreviated as the K-loop), and was designed and inserted into two different regions of the protein bacterial alkaline phosphatase (BAP). MTG-mediated selective labeling of a lysine residue in the K-loop was achieved with a functional Gln-donor substrate. Internal protein labeling in the vicinity of the active site of BAP (residues 91-93) markedly decreased the activity of the enzyme. Conversely, insertion of the K-loop at a site distal from the active site (residues 219-221) afforded site-specific and covalent internal protein labeling without impairing the activity of the enzyme.. |
| 295. |
Josui Shimoda, Tatsuo Maruyama, Momoko Kitaoka, Noriho Kamiya, Masahiro Goto, DNA-enzyme conjugate with a weak inhibitor that can specifically detect thrombin in a homogeneous medium, ANALYTICAL BIOCHEMISTRY, 10.1016/j.ab.2011.02.035, 414, 1, 103-108, 2011.07, We present the DNA-assisted control of enzymatic activity for the detection of a target protein using a new type of DNA-enzyme conjugate. The conjugate is composed of an enzyme inhibitor to regulate enzyme activity and a DNA aptamer to be responsive toward the analyte protein. Glutathione S-transferase (GST) and thrombin were selected as a model enzyme and an analyte protein. A hexahistidine tag was genetically attached to the C terminus of the GST, and the 5' end of an oligonucleotide was conjugated with nitrilotriacetic acid (NTA) for the site-specific conjugation of the DNA with the GST based on a Ni(2+) complex interaction. We found that fluorescein acted as a weak inhibitor of GST and succeeded in the regulation of GST activity by increasing the local concentration of the weak inhibitor by the hybridization of a 3'-end fluorescein-modified DNA. The catalytic activity of the DNA aptamer-enzyme conjugate showed a dose-dependent response to thrombin, indicating that the GST activity was clearly recovered by the binding of the DNA aptamer to thrombin. The current system enables the sensitive and specific detection of thrombin simply by measuring the enzymatic activity in a homogeneous medium. (C) 2011 Elsevier Inc. All rights reserved.. |
| 296. |
Yutaro Mori, Masahiro Goto, Noriho Kamiya, Transglutaminase-mediated internal protein labeling with a designed peptide loop, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 10.1016/j.bbrc.2011.06.073, 410, 4, 829-833, 2011.07, Post-translational internal protein labeling was explored through the insertion of a 13-mer peptidyl loop specifically recognized by microbial transglutaminase (MTG). The peptidyl loop included one lysine residue (abbreviated as the K-loop), and was designed and inserted into two different regions of the protein bacterial alkaline phosphatase (BAP). MTG-mediated selective labeling of a lysine residue in the K-loop was achieved with a functional Gln-donor substrate. Internal protein labeling in the vicinity of the active site of BAP (residues 91-93) markedly decreased the activity of the enzyme. Conversely, insertion of the K-loop at a site distal from the active site (residues 219-221) afforded site-specific and covalent internal protein labeling without impairing the activity of the enzyme. (C) 2011 Elsevier Inc. All rights reserved.. |
| 297. |
Kousuke Moriyama, Kyunga Sung, Masahiro Goto, Noriho Kamiya, Immobilization of alkaline phosphatase on magnetic particles by site-specific and covalent cross-linking catalyzed by microbial transglutaminase, Journal of Bioscience and Bioengineering, 10.1016/j.jbiosc.2011.02.002, 111, 6, 650-653, 2011.06, Bacterial alkaline phosphatase (BAP) was site-specifically and covalently immobilized on magnetic particles (MPs) using the enzymatic reaction of microbial transglutaminase (MTG). Immobilization efficiency was affected by the chemical surface treatment of MPs and immobilized BAP exhibited more than 90% of the initial activity after 10 rounds of recycling.. |
| 298. |
Kousuke Moriyama, Kyunga Sung, Masahiro Goto, Noriho Kamiya, Immobilization of alkaline phosphatase on magnetic particles by site-specific and covalent cross-linking catalyzed by microbial transglutaminase, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 10.1016/j.jbiosc.2011.02.002, 111, 6, 650-653, 2011.06, Bacterial alkaline phosphatase (BAP) was site-specifically and covalently immobilized on magnetic particles (MPs) using the enzymatic reaction of microbial transglutaminase (MTG). Immobilization efficiency was affected by the chemical surface treatment of MPs and immobilized BAP exhibited more than 90% of the initial activity after 10 rounds of recycling. (C) 2011, The Society for Biotechnology, Japan. All rights reserved.. |
| 299. |
T. Niide, M. Goto, N. Kamiya, Biocatalytic synthesis of gold nanoparticles with cofactor regeneration in recombinant Escherichia coli cells, Chem. Commun., 47, 7350-7352, 2011.05. |
| 300. |
Y. Mori, M. Goto, N. Kamiya, Transglutaminase-mediated internal protein labeling with a designed peptide loop, Biochem. Biophys. Res. Commun., 410, 829-833, 2011.05. |
| 301. |
Y. Mori, K. Minamihata, H. Abe, M. Goto, N. Kamiya, Protein assemblies by site-specific avidin-biotin interactions, Org. Biomol. Chem., 9, 5641-5644, 2011.05. |
| 302. |
Momoko Kitaoka, Yukito Tsuruda, Yukari Tanaka, Masahiro Goto, Masayuki Mitsumori, Kounosuke Hayashi, Yoshiyuki Hiraishi, Katsuyuki Miyawaki, Sumihare Noji, Noriho Kamiya, Transglutaminase-mediated synthesis of a DNA-(Enzyme)n probe for highly sensitive DNA detection, Chemistry - A European Journal, 10.1002/chem.201003744, 17, 19, 5387-5392, 2011.05, A new synthetic strategy for DNA-enzyme conjugates with a novel architecture was explored using a natural cross-linking catalyst, microbial transglutaminase (MTG). A glutamine-donor substrate peptide of MTG was introduced at the 5-position on the pyrimidine of deoxyuridine triphosphate to prepare a DNA strand with multiple glutamine-donor sites by polymerase chain reaction (PCR). A substrate peptide that contained an MTG-reactive lysine residue was fused to the N terminus of a thermostable alkaline phoshatase from Pyrococcus furiosus (PfuAP) by genetic engineering. By combining enzymatically the substrate moieties of MTG introduced to the DNA template and the recombinant enzyme, a DNA-(enzyme)n conjugate with 1:n stoichiometry was successfully obtained. The enzyme/DNA ratio of the conjugate increased as the benzyloxycarbonyl-L-glutaminylglycine (Z-QG) moiety increased in the DNA template. The potential utility of the new conjugate decorated with signaling enzymes was validated in a dot blot hybridization assay. The DNA-(enzyme) n probe could clearly detect 104 copies of the target nucleic acid with the complementary sequence under harsh hybridization conditions, thereby enabling a simple detection procedure without cumbersome bound/free processes associated with a conventional hapten-antibody reaction-based DNA-detection system. DNA detector: A glutamine-modified DNA probe was synthesized by the polymerase chain reaction by using a glutamine-modified 2′-deoxyuridine 5′-triphosphate analogue as a substrate of DNA polymerase. The DNA-(alkaline phosphatase)n conjugate probe was highly sensitive and could directly visualize the target DNA bound on a membrane immediately after hybridization (see graphic).. |
| 303. |
Momoko Kitaoka, Yukito Tsuruda, Yukari Tanaka, Masahiro Goto, Masayuki Mitsumori, Kounosuke Hayashi, Yoshiyuki Hiraishi, Katsuyuki Miyawaki, Sumihare Noji, Noriho Kamiya, Transglutaminase-Mediated Synthesis of a DNA-(Enzyme)(n) Probe for Highly Sensitive DNA Detection, CHEMISTRY-A EUROPEAN JOURNAL, 10.1002/chem.201003744, 17, 19, 5387-5392, 2011.05, A new synthetic strategy for DNA-enzyme conjugates with a novel architecture was explored using a natural cross-linking catalyst, microbial transglutaminase (MTG). A glutamine-donor substrate peptide of MTG was introduced at the 5-position on the pyrimidine of deoxyuridine triphosphate to prepare a DNA strand with multiple glutamine-donor sites by polymerase chain reaction (PCR). A substrate peptide that contained an MTG-reactive lysine residue was fused to the N terminus of a thermostable alkaline phoshatase from Pyrococcus furiosus (PfuAP) by genetic engineering. By combining enzymatically the substrate moieties of MTG introduced to the DNA template and the recombinant enzyme, a DNA-(enzyme)(n) conjugate with 1:n stoichiometry was successfully obtained. The enzyme/DNA ratio of the conjugate increased as the benzyloxycarbonyl-L-glutaminylglycine (ZQG) moiety increased in the DNA template. The potential utility of the new conjugate decorated with signaling enzymes was validated in a dot blot hybridization assay. The DNA-(enzyme)(n) probe could clearly detect 10(4) copies of the target nucleic acid with the complementary sequence under harsh hybridization conditions, thereby enabling a simple detection procedure without cumbersome bound/free processes associated with a conventional hapten-antibody reaction-based DNA-detection system.. |
| 304. |
K. Minamihata, M. Goto, N. Kamiya, Site-specific protein cross-linking by peroxidase-catalyzed activation of a tyrosine-containing peptide tag, Bioconjugate Chem., 22, 74-81, 2011.04. |
| 305. |
K. Moriyama, K. Sung, M. Goto, N. Kamiya, Immobilization of alkaline phosphatase on magnetic particles by site-specific and covalent cross-linking catalyzed by microbial transglutaminase, J. Biosci. Bioeng., 111, 650-653, 2011.04. |
| 306. |
M. Sakono, K. Motomura, T. Maruyama, N. Kamiya, M. Goto, Alpha casein micelles show not only molecular chaperone-like aggregation inhibition properties but also protein refolding activity from the denatured state, Biochem. Biophys. Res. Commun., 404, 494-497, 2011.04. |
| 307. |
Momoko Kitaoka, Yukari Tanaka, Yutaka Tada, Masahiro Goto, Katsuyuki Miyawaki, Sumihare Noji, Noriho Kamiya, Conjugation of enzymes on RNA probes through Cu(I) catalyzed alkyne-azide cycloaddition, Biotechnology Journal, 10.1002/biot.201000249, 6, 4, 470-476, 2011.04, Northern and Southern blots are the most commonly used techniques for the confirmation of presence and expression of target genes. Molecular tools available for this purpose include radioisotope-, enzyme- and hapten-labeled nucleic acid probes. In particular, the use of enzyme-labeled probes are easy and safe, and do not require bound/free processes after hybridization associated with an antibody-based detection system. However, there are few approaches that enable the post-transcriptional modification of RNA with enzymes or proteins. In this study, we applied the Cu(I)-catalyzed [3 + 2] azide-alkyne cycloaddition (CuAAC) reaction to the labeling of an RNA strand with enzymes. The C-5 position of UTP was modified with an alkyne group and alkyne-bearing RNA was prepared by in vitro transcription using T7 RNA polymerase. Surface amino groups of bacterial alkaline phosphatase (BAP) were randomly derivatized with azide groups at different modification ratios. The CuAAC reaction occurred selectively between the alkyne-modified RNA and the azide-modified enzyme. The RNA probe conjugated with BAP using this technique could detect a specific RNA by dot blot northern hybridization.. |
| 308. |
Momoko Kitaoka, Yukari Tanaka, Yutaka Tada, Masahiro Goto, Katsuyuki Miyawaki, Sumihare Noji, Noriho Kamiya, Conjugation of enzymes on RNA probes through Cu(I) catalyzed alkyne-azide cycloaddition, BIOTECHNOLOGY JOURNAL, 10.1002/biot.201000249, 6, 4, 470-476, 2011.04, Northern and Southern blots are the most commonly used techniques for the confirmation of presence and expression of target genes. Molecular tools available for this purpose include radioisotope-, enzyme-and hapten-labeled nucleic acid probes. In particular, the use of enzyme-labeled probes are easy and safe, and do not require bound/free processes after hybridization associated with an antibody-based detection system. However, there are few approaches that enable the post-transcriptional modification of RNA with enzymes or proteins. In this study, we applied the Cu(I)-catalyzed [3 + 2] azide-alkyne cycloaddition (CuAAC) reaction to the labeling of an RNA strand with enzymes. The C-5 position of UTP was modified with an alkyne group and alkyne-bearing RNA was prepared by in vitro transcription using T7 RNA polymerase. Surface amino groups of bacterial alkaline phosphatase (BAP) were randomly derivatized with azide groups at different modification ratios. The CuAAC reaction occurred selectively between the alkyne-modified RNA and the azide-modified enzyme. The RNA probe conjugated with BAP using this technique could detect a specific RNA by dot blot northern hybridization.. |
| 309. |
M. Kitaoka, Y. Tsuruda, Y. Tanaka, M. Goto, M. Mitsumori, K. Hayashi, Y. Hiraishi, K. Miyawaki, S. Noji, N. Kamiya, Transglutaminase-mediated synthesis of a novel DNA-(enzyme)n probe for highly sensitive DNA detection, Chem. Eur. J., 19, 5387-5392, 2011.03, 生体内ではほとんどのタンパク質が何らかの翻訳後修飾を受けることに着目し、翻訳後修飾過程で働く酵素の基質特異性を利用すれば、狙った部位でタンパク質を修飾できると考えた。従来の有機化学的手法では、タンパク質の狙った部位を選択的に修飾するのは極めて困難であった。そこでラベル化したい有機分子内に基質となる部位（酵素の認識部位）を設計し、これを糊代として利用することで、糊代選択的な生体分子の連結・修飾法を確立した。
既往の方法ではDNAとタンパク質を１：１で連結することしかできなかったが、DNAを構成する分子を酵素の基質となるように設計し、DNA-(タンパク質)n型の新規ハイブリッド分子を高収率で得る技術を確立し、新たな遺伝子検出システムを提案した。. |
| 310. |
D. Pissuwan, K. Nose, R. Kurihara, K. Kaneko, Y. Tahara, N. Kamiya, M. Goto, Y. Katayama, T. Niidome, A solid-in-oil dispersion of gold nanorods can enhance trandermal protein delivery and skin vaccination, small, 7, 215-220, 2011.02. |
| 311. |
チロシン酸化反応を用いた異種タンパク質架橋体の酵素合成. |
| 312. |
T. Mouri, N. Kamiya,* M. Goto, New strategy to enhance catalytic performance of Escherichia coli whole cell biocatalysts harboring P450cam mutants, Biochem. Eng. J., 53, 229-233, 2011.01. |
| 313. |
Dakrong Pissuwan, Keisuke Nose, Ryohsuke Kurihara, Kenji Kaneko, Yoshiro Tahara, Noriho Kamiya, Masahiro Goto, Yoshiki Katayama, Takuro Niidome, A solid-in-oil dispersion of gold nanorods can enhance transdermal protein delivery and skin vaccination, Small, 10.1002/smll.201001394, 7, 2, 215-220, 2011.01, A high-molecular-weight model protein, ovalbumin (OVA), can be delivered through the skin and induce an immune response in mice by a solid-in-oil dispersion of gold nanorods after irradiation with near-infrared light.. |
| 314. |
Masafumi Sakono, Konomi Motomura, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Alpha casein micelles show not only molecular chaperone-like aggregation inhibition properties but also protein refolding activity from the denatured state, Biochemical and Biophysical Research Communications, 10.1016/j.bbrc.2010.12.009, 404, 1, 494-497, 2011.01, Casein micelles are a major component of milk proteins. It is well known that casein micelles show chaperone-like activity such as inhibition of protein aggregation and stabilization of proteins. In this study, it was revealed that casein micelles also possess a high refolding activity for denatured proteins. A buffer containing caseins exhibited higher refolding activity for denatured bovine carbonic anhydrase than buffers including other proteins. In particular, a buffer containing α-casein showed about a twofold higher refolding activity compared with absence of α-casein. Casein properties of surface hydrophobicity, a flexible structure and assembly formation are thought to contribute to this high refolding activity. Our results indicate that casein micelles stabilize milk proteins by both chaperone-like activity and refolding properties.. |
| 315. |
Noriho Kamiya, Hiroki Abe, New fluorescent substrates of microbial transglutaminase and its application to peptide tag-directed covalent protein labeling, Methods in Molecular Biology, 10.1007/978-1-61779-151-2_7, 81-94, 2011.01, Transglutaminase (TGase) is an enzyme that catalyzes the post-translational covalent cross-linking of Gln-and Lys-containing peptides and/or proteins according to its substrate specificity. We have recently designed a variety of Gln-donor fluorescent substrates of microbial transglutaminase (MTG) from Streptomyces mobaraensis and evaluated their potential use in MTG-mediated covalent protein labeling. The newly designed substrates are based on the relatively broad substrate recognition of MTG for the substitution of the N-terminal group of a conventional TGase substrate, benzyloxycarbonyl-l-glutaminylglycine (Z-QG). It is revealed that MTG is capable of accepting a diverse range of fluorophores in place of the N-terminal moiety of Z-QG when linked via a suitable linker. Here, we show the potential utility of a new fluorescent substrate for peptide tag-directed covalent protein labeling by employing fluorescein-4-isothiocyanate-β-Ala-QG as a model Gln-donor substrate for MTG.. |
| 316. |
Tsuyoshi Mouri, Noriho Kamiya, Masahiro Goto, New strategy to enhance catalytic performance of Escherichia coli whole cell biocatalysts harboring P450cam mutants, Biochemical Engineering Journal, 10.1016/j.bej.2010.09.016, 53, 2, 229-233, 2011.01, Cytochrome P450cam mutants (Y96F and F87W-Y96F) can catalyze the oxidation of dichlorobenzene and other aromatic compound that wild type P450cam cannot catalyze. The expression of the single mutant Y96F in an Escherichia coli host resulted in indigo formation (approximately 23mg/l), of which productivity was comparable to the highest reported so far using other P450cam mutant cultures. On the other hand, recombinant E. coli that harbored F87W-Y96F exhibited little activity for indigo production. However, co-expression of E. coli glycerol dehydrogenase (GLD) with F87W-Y96F triggered a marked increase in the productivity of indigo (approximately 20mg/l) without external glycerol addition. These results indicate the potential of introducing GLD-mediated NADH regeneration that utilizes endogenous NAD+ and glycerol in E. coli to enhance markedly the catalytic performance of a low-activity recombinant P450cam system. The present E. coli whole-cell biocatalysts might be applicable to practical microbial indigo production.. |
| 317. |
Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Site-specific protein cross-linking by peroxidase-catalyzed activation of a tyrosine-containing peptide tag, Bioconjugate Chemistry, 10.1021/bc1003982, 22, 1, 74-81, 2011.01, Protein modification methods represent fundamental techniques that are applicable in many fields. In this study, a site-specific protein cross-linking based on the oxidative tyrosine coupling reaction was demonstrated. In the presence of horseradish peroxidase (HRP) and H2O2, tyrosine residues undergo one-electron oxidation reactions and form radicals in their phenolic moieties, and these species subsequently react with each other to form dimers or further react to generate polymers. Here, a peptide-tag containing a tyrosine residue(s) (Y-tag, of which the amino acid sequences were either GGGGY or GGYYY) was genetically introduced at the C-terminus of a model protein, Escherichia coli alkaline phosphatase (BAP). Following the incubation of recombinant BAPs with HRP and H2O2, Y-tagged BAPs were efficiently cross-linked with each other, whereas wild-type BAP did not undergo cross-linking, indicating that the tyrosine residues in the Y-tags were recognized by HRP as the substrates. To determine the site-specificity of the cross-linking reaction, the Y-tag was selectively removed by thrombin digestion. The resultant BAP without the Y-tag showed no reactivity in the presence of HRP and H2O2. Conversely, Y-tagged BAPs cross-linked by HRP treatment were almost completely digested into monomeric BAP units following incubation with the protease. Moreover, cross-linked Y-tagged BAPs retained ∼95% of their native enzymatic activity. These results show that HRP catalyzed the site-specific cross-linking of BAPs through tyrosine residues positioned in the C-terminal Y-tag. The site-selective enzymatic oxidative tyrosine coupling reaction should offer a practical option for site-specific and covalent protein modifications.. |
| 318. |
Dakrong Pissuwan, Keisuke Nose, Ryohsuke Kurihara, Kenji Kaneko, Yoshiro Tahara, Noriho Kamiya, Masahiro Goto, Yoshiki Katayama, Takuro Niidome, A Solid-in-Oil Dispersion of Gold Nanorods Can Enhance Transdermal Protein Delivery and Skin Vaccination, SMALL, 10.1002/smll.201001394, 7, 2, 215-220, 2011.01, A high-molecular-weight model protein, ovalbumin (OVA), can be delivered through the skin and induce an immune response in mice by a solid-in-oil dispersion of gold nanorods after irradiation with near-infrared light. © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.. |
| 319. |
Masafumi Sakono, Konomi Motomura, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Alpha casein micelles show not only molecular chaperone-like aggregation inhibition properties but also protein refolding activity from the denatured state, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 10.1016/j.bbrc.2010.12.009, 404, 1, 494-497, 2011.01, Casein micelles are a major component of milk proteins. It is well known that casein micelles show chaperone-like activity such as inhibition of protein aggregation and stabilization of proteins. In this study, it was revealed that casein micelles also possess a high refolding activity for denatured proteins. A buffer containing caseins exhibited higher refolding activity for denatured bovine carbonic anhydrase than buffers including other proteins. In particular, a buffer containing alpha-casein showed about a twofold higher refolding activity compared with absence of alpha-casein. Casein properties of surface hydrophobicity, a flexible structure and assembly formation are thought to contribute to this high refolding activity. Our results indicate that casein micelles stabilize milk proteins by both chaperone-like activity and refolding properties. (C) 2010 Elsevier Inc. All rights reserved.. |
| 320. |
Tsuyoshi Mouri, Noriho Kamiya, Masahiro Goto, New strategy to enhance catalytic performance of Escherichia coli whole cell biocatalysts harboring P450cam mutants, BIOCHEMICAL ENGINEERING JOURNAL, 10.1016/j.bej.2010.09.016, 53, 2, 229-233, 2011.01, Cytochrome P450cam mutants (Y96F and F87W-Y96F) can catalyze the oxidation of dichlorobenzene and other aromatic compound that wild type P450cam cannot catalyze. The expression of the single mutant Y96F in an Escherichia coli host resulted in indigo formation (approximately 23 mg/l), of which productivity was comparable to the highest reported so far using other P450cam mutant cultures. On the other hand, recombinant E. coli that harbored F87W-Y96F exhibited little activity for indigo production. However, co-expression of E. coli glycerol dehydrogenase (GLD) with F87W-Y96F triggered a marked increase in the productivity of indigo (approximately 20 mg/l) without external glycerol addition. These results indicate the potential of introducing GLD-mediated NADH regeneration that utilizes endogenous NAD(+) and glycerol in E. coli to enhance markedly the catalytic performance of a low-activity recombinant P450cam system. The present E. coli whole-cell biocatalysts might be applicable to practical microbial indigo production. (C) 2010 Elsevier B.V. All rights reserved.. |
| 321. |
Kosuke Minamihata, Masahiro Goto, Noriho Kamiya, Site-Specific Protein Cross-Linking by Peroxidase-Catalyzed Activation of a Tyrosine-Containing Peptide Tag, BIOCONJUGATE CHEMISTRY, 10.1021/bc1003982, 22, 1, 74-81, 2011.01, Protein modification methods represent fundamental techniques that are applicable in many fields. In this study, a site-specific protein cross-linking based on the oxidative tyrosine coupling reaction was demonstrated. In the presence of horseradish peroxidase (HRP) and H(2)O(2), tyrosine residues undergo one-electron oxidation reactions and form radicals in their phenolic moieties, and these species subsequently react with each other to form dimers or further react to generate polymers. Here, a peptide-tag containing a tyrosine residue(s) (Y-tag, of which the amino acid sequences were either GGGGY or GGYYY) was genetically introduced at the C-terminus of a model protein, Escherichia coli alkaline phosphatase (BAP). Following the incubation of recombinant BAPs with HRP and H(2)O(2), Y-tagged BAPs were efficiently cross-linked with each other, whereas wild-type BAP did not undergo cross-linking, indicating that the tyrosine residues in the Y-tags were recognized by HRP as the substrates. To determine the site-specificity of the cross-linking reaction, the Y-tag was selectively removed by thrombin digestion. The resultant BAP without the Y-tag showed no reactivity in the presence of HRP and H(2)O(2). Conversely, Y-tagged BAPs cross-linked by HRP treatment were almost completely digested into monomeric BAP units following incubation with the protease. Moreover, cross-linked Y-tagged BAPs retained similar to 95% of their native enzymatic activity. These results show that HRP catalyzed the site-specific cross-linking of BAPs through tyrosine residues positioned in the C-terminal Y-tag. The site-selective enzymatic oxidative tyrosine coupling reaction should offer a practical option for site-specific and covalent protein modifications.. |
| 322. |
Fukiko Kubota, Yousuke Shimobori, Yuzo Baba, Yusuke Koyanagi, Kojiro Shimojo, Noiho Kamiya, Masahiro Goto, Application of ionic liquids to extraction separation of rare earth metals with an effective diglycol amic acid extractant, JOURNAL OF CHEMICAL ENGINEERING OF JAPAN, 10.1252/jcej.11we005, 44, 5, 307-312, 2011, The application of ionic liquids as alternatives to conventional organic solvents in extraction processes has been actively investigated. A crucial step towards the practical use of ionic liquids is the development of extractants that work effectively within these new media. In the present study, the extraction separation of rare earth metals into ionic liquids, 1-butyl, 1-octyl, and 1-dodecyl-3-methylimidazolium bis(trifluoromethanesulfonyl)imide ([Cnmim][Tf2N], n=4, 8, 12), was performed using a novel extractant, N,N-dioctyldiglycol amic acid (DODGAA). Quantitative extraction of metal ions such as Y3+ and Eu3+ was selectively achieved in the presence of the base metal ion Zn2+, which was not extracted at all under the present experimental conditions. The extraction efficiency was enhanced for the shorter-alkyl-chain imidazolium ionic liquid [C4mim][Tf2N] compared to that for a conventional organic solvent system. Extraction mechanism studies elucidated that the metal extraction proceeds via proton exchange reactions between DODGAA and the metal ions in the ionic liquid (the same mechanism as in the conventional organic solvent). The stripping reaction, or recovery, of the metal ions from the extracting phase was readily accomplished with an acid solution such as nitric acid.. |
| 323. |
Nobuiro Ishida, Satoshi Katahira, Wataru Tokuhara, Yoshiyuki Noritake, Noriho Kamiya, Kazunori Nakashima, Chiaki Ogino, Akihiko Kondo, Development of ionic liquid-based consolidated bioprocessing (I-CBP) for bioethanol production, Sustainable Engineering Forum: Core Programming Topic at the 2011 AIChE Annual Meeting
Sustainable Engineering Forum
Core Programming Topic at the 2011 AIChE Annual Meeting, 2011. |
| 324. |
Yuzo Baba, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Selective recovery of dysprosium and neodymium ions by a supported liquid membrane based on ionic liquids, Solvent Extraction Research and Development, 10.15261/serdj.18.193, 18, 193-198, 2011, Selective permeation of rare earth metal ions (Dy and Nd) against ferric ion (Fe) was conducted by an ionic liquid-based supported liquid membrane (SLM). A novel extractant N,Ndioctyldiglycol amic acid (DODGAA) was employed as the mobile carrier because the DODGAA is readily soluble in ionic liquids such as l-octyl-3-methylimidazolium bis(trifluoromethanesulfonyl)imide ([Cgmim][Tf 2N]) and shows good selectivity to the rare earth metal ions and efficient stripping behavior. Quantitative transport of Dy and Nd was successfully achieved through the SLM impregnated with the [Cgmim][Tf 2N] containing DODGAA. Ferric ions were transported at the 10 % level, and Dy and Nd were selectively recovered to the receiving phase. The results suggested a potential use of the SLM system proposed in this study for the recovery of the rare earth metals from leaching magnet scrap which produces leach liquor containing a large amount of Fe.. |
| 325. |
Muhammad Moniruzzaman, Noriho Kamiya, Masahiro Goto, Ionic liquid based microemulsion with pharmaceutically accepted components
Formulation and potential applications, Journal of Colloid And Interface Science, 10.1016/j.jcis.2010.08.035, 352, 1, 136-142, 2010.12, In this paper, we report a novel ionic liquid-in-oil (IL/o) microemulsion which is able to dissolve pharmaceuticals that are insoluble or sparingly soluble in water and most of pharmaceutical grade organic liquids. Towards this approach, the nanometer-sized ionic liquid droplets in isopropyl myristate (IPM) were formed with a blend of nonionic surfactants, polyoxyethylene sorbitan monooleate (Tween-80), and sorbitan laurate (Span-20). A set of ionic liquids (ILs) was tested as a dispersed phase, and it was observed that ILs possessing coordinating anions which are strong hydrogen bond acceptor were most effective in forming microemulsion droplets. The possible formation mechanism was also studied. Ternary phase behavior study clearly indicated the formation of optically transparent and thermodynamically stable microemulsions with a wide range of IL content. The shape, size and size distribution of the aggregates in microemulsions were characterized using dynamic light scattering (DLS), which demonstrated the formation of spherical micelles in the range of 8-34 nm. In order to explore the use of newly developed microemulsion as a potential drug carrier, we have investigated the solubility of some drug molecules (e.g., acyclovir, methotrexate and 1-[(5-(p-nitrophenyl) furfurylidene) amino] hydantoin sodium) that are insoluble or sparingly soluble in most of the conventional solvents. Very significantly, the solubility studies indicated a high degree of solubilization of such drugs in IL microemulsions. We believe that this microemulsion formed with ILs having the unique physical, chemical and biological properties may offer novel opportunities to develop a potential drug delivery carrier for poorly soluble drugs molecules.. |
| 326. |
Yoshiro Tahara, Kenichi Namatsu, Noriho Kamiya, Masayori Hagimori, Seitaro Kamiya, Masayuki Arakawa, Masahiro Goto, Transcutaneous immunization by a solid-in-oil nanodispersion, Chemical Communications, 10.1039/c0cc03600e, 46, 48, 9200-9202, 2010.12, We have successfully achieved transcutaneous immunization without the use of any skin pre-treatment or immune-stimulant adjuvant by applying a solid-in-oil (S/O) nanodispersion: an oil-based nanodispersion of antigens coated with hydrophobic surfactant molecules. This finding indicates that the S/O nanodispersion has great promise for effective transcutaneous vaccination.. |
| 327. |
Muhammad Moniruzzaman, Noriho Kamiya, Masahiro Goto, Ionic liquid based microemulsion with pharmaceutically accepted components: Formulation and potential applications, JOURNAL OF COLLOID AND INTERFACE SCIENCE, 10.1016/j.jcis.2010.08.035, 352, 1, 136-142, 2010.12, In this paper, we report a novel ionic liquid-in-oil (IL/o) microemulsion which is able to dissolve pharmaceuticals that are insoluble or sparingly soluble in water and most of pharmaceutical grade organic liquids. Towards this approach, the nanometer-sized ionic liquid droplets in isopropyl myristate (IPM) were formed with a blend of nonionic surfactants, polyoxyethylene sorbitan monooleate (Tween-80), and sorbitan laurate (Span-20). A set of ionic liquids (ILs) was tested as a dispersed phase, and it was observed that ILs possessing coordinating anions which are strong hydrogen bond acceptor were most effective in forming microemulsion droplets. The possible formation mechanism was also studied. Ternary phase behavior study clearly indicated the formation of optically transparent and thermodynamically stable microemulsions with a wide range of IL content. The shape, size and size distribution of the aggregates in microemulsions were characterized using dynamic light scattering (DLS), which demonstrated the formation of spherical micelles in the range of 8-34 nm. In order to explore the use of newly developed microemulsion as a potential drug carrier, we have investigated the solubility of some drug molecules (e.g., acyclovir, methotrexate and 1-[(5-(p-nitrophenyl) furfurylidene) amino] hydantoin sodium) that are insoluble or sparingly soluble in most of the conventional solvents. Very significantly, the solubility studies indicated a high degree of solubilization of such drugs in IL microemulsions. We believe that this microemulsion formed with ILs having the unique physical, chemical and biological properties may offer novel opportunities to develop a potential drug delivery carrier for poorly soluble drugs molecules. (C) 2010 Elsevier Inc. All rights reserved.. |
| 328. |
Muhammad Moniruzzaman, Miki Tamura, Yoshiro Tahara, Noriho Kamiya, Masahiro Goto, Ionic liquid-in-oil microemulsion as a potential carrier of sparingly soluble drug
Characterization and cytotoxicity evaluation, International Journal of Pharmaceutics, 10.1016/j.ijpharm.2010.08.034, 400, 1-2, 243-250, 2010.11, Pharmaceutical industries have posed challenges in the topical and transdermal administration of drugs which are poorly soluble or insoluble in water and most of organic solvents. In an approach to overcome this limitation, ionic liquid-in-oil (IL/o) microemulsions (MEs) were employed to increase the solubility of a sparingly soluble drug to enhance its topical and transdermal delivery. The formulation of MEs was composed of a blend of nonionic surfactants, polyoxyethylene sorbitan monooleate (Tween-80) and sorbitan laurate (Span-20), isopropyl myristate (IPM) as an oil phase, and IL [C1mim] [(CH3O)2PO2] (dimethylimidazolium dimethylphosphate) as a pseudophase. Among various weight ratios of Tween-80 to Span-20 investigated in the ME systems, the ratio 3:2 showed excellent solubility and skin permeation enhancing effect for acyclovir (ACV) used as a model sparingly soluble drug. The size and size distribution of the ME droplets with and without drug were determined by dynamic light scattering. The permeability study of ACV incorporated in IL droplets as well as other formulations was performed into and across the Yucatan micropig (YMP) porcine skin, and the use of IL/o MEs has been shown to dramatically increase ACV administration. Finally, the cytotoxicity of the new carrier was evaluated in vitro using the reconstructed human epidermal model LabCyte™ EPI-MODEL12. It was found that the cell viability of IL/o MEs containing 4wt% IL was over 80% compared to Dulbecco's Phosphate-Buffered Salines, indicating low cytotoxicity of the carrier. Taken together these results, it can be assumed that IL-assisted nonaqueous ME could serve as a versatile and efficient nanodelivery system for insoluble or sparingly soluble drug molecules that require solubilizing agents for delivery.. |
| 329. |
Muhammad Moniruzzaman, Miki Tamura, Yoshiro Tahara, Noriho Kamiya, Masahiro Goto, Ionic liquid-in-oil microemulsion as a potential carrier of sparingly soluble drug: Characterization and cytotoxicity evaluation, INTERNATIONAL JOURNAL OF PHARMACEUTICS, 10.1016/j.ijpharm.2010.08.034, 400, 1-2, 243-250, 2010.11, Pharmaceutical industries have posed challenges in the topical and transdermal administration of drugs which are poorly soluble or insoluble in water and most of organic solvents. In an approach to overcome this limitation, ionic liquid-in-oil (IL/o) microemulsions (MEs) were employed to increase the solubility of a sparingly soluble drug to enhance its topical and transdermal delivery. The formulation of MEs was composed of a blend of nonionic surfactants, polyoxyethylene sorbitan monooleate (Tween-80) and sorbitan laurate (Span-20), isopropyl myristate (IPM) as an oil phase, and IL [C(i)mim],[(CH(3)O)(2)PO(2)] (dimethylimidazolium dimethylphosphate) as a pseudophase. Among various weight ratios of Tween-80 to Span-20 investigated in the ME systems, the ratio 3:2 showed excellent solubility and skin permeation enhancing effect for acyclovir (ACV) used as a model sparingly soluble drug. The size and size distribution of the ME droplets with and without drug were determined by dynamic light scattering. The permeability study of ACV incorporated in IL droplets as well as other formulations was performed into and across the Yucatan micropig (YMP) porcine skin, and the use of IL/o MEs has been shown to dramatically increase ACV administration. Finally, the cytotoxicity of the new carrier was evaluated in vitro using the reconstructed human epidermal model LabCyte (TM) EPI-MODEL12. It was found that the cell viability of IL/o MEs containing 4 wt% IL was over 80% compared to Dulbecco's Phosphate-Buffered Salines, indicating low cytotoxicity of the carrier. Taken together these results, it can be assumed that IL-assisted nonaqueous ME could serve as a versatile and efficient nanodelivery system for insoluble or sparingly soluble drug molecules that require solubilizing agents for delivery. (C) 2010 Elsevier B.V. All rights reserved.. |
| 330. |
新規カチオン性ダブルコーティング型キャリアによる遺伝子デリバリー. |
| 331. |
Hiroki Abe, Masahiro Goto, Noriho Kamiya, Enzymatic single-step preparation of multifunctional proteins, Chemical Communications, 10.1039/c0cc02133d, 46, 38, 7160-7162, 2010.10, A small glutamine donor substrate was designed for single-step and site-specific protein multifunctionalization catalyzed by microbial transglutaminase.. |
| 332. |
部位特異的タンパク質修飾のための高反応性トランスグルタミナーゼ基質ペプチド配列の探索. |
| 333. |
Yukiko Shimada, Teppei Niide, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Selective separation of precious metals using biomass materials, kagaku kogaku ronbunshu, 10.1252/kakoronbunshu.36.255, 36, 4, 255-258, 2010.07, Proteins are known to exhibit specific interactions with various metalions. In this study, we investigated two types of protein-rich biomass, chicken feather and hen eggshell membrane, as adsorbents for separation of precious metal ions. Precious metal ions such as Au(III), Pd(II) and Pt(IV) were selectively adsorbed on the biomass in the presence of various transition metal ions. The adsorption capacity of eggshell membrane was higher than that of feather. Adsorption of Au(III) on the eggshell membrane was prominent at a pH range below 5. The adsorption behavior was explained by the complexation of Au(III) ion with proteins in the eggshell membrane and it was affected by Au(III) ion species in the feed aqueous solutions.. |
| 334. |
ファージディスプレイ法を用いた部位特異的タンパク質修飾のためのペプチドタグの探索. |
| 335. |
Yoshiro TAHARA, Kenichi NAMATSU, Noriho KAMIYA, Masayori HAGIMORI, Shinji KAMIYA, Masayuki ARAKAWA, Masahiro GOTO, Transcutaneous immunization by a solid-in-oil nanodispersion, Chem. Commun., 46, 9200-9202, 2010.06. |
| 336. |
K. Sung, N. Kamiya,* N. Kawata, S. Kamiya and M. Goto, Functional glass surface displaying a glutamyl donor substrate for transglutaminase-mediated protein immobilization, Biotechnol. J., 46, 456-462, 2010.06. |
| 337. |
Muhammad Moniruzzaman, Noriho Kamiya, Masahiro Goto, Activation and stabilization of enzymes in ionic liquids, Organic and Biomolecular Chemistry, 10.1039/b926130c, 8, 13, 2887-2899, 2010.06, As environmentally benign "green" solvents, room temperature ionic liquids (ILs) have been used as solvents or (co)solvents in biocatalytic reactions and processes for a decade. The technological utility of enzymes can be enhanced greatly by their use in ionic liquids (ILs) rather than in conventional organic solvents or in their natural aqueous reaction media. In fact, the combination of green properties and unique tailor-made physicochemical properties make ILs excellent non-aqueous solvents for enzymatic catalysis with numerous advantages over other solvents, including high conversion rates, high selectivity, better enzyme stability, as well as better recoverability and recyclability. However, in many cases, particularly in hydrophilic ILs, enzymes show relative instability and/or lower activity compared with conventional solvents. To improve the enzyme activity as well as stability in ILs, various attempts have been made by modifying the form of the enzymes. Examples are enzyme immobilization onto support materials via adsorption or multipoint attachment, lyophilization in the presence of stabilizing agents, chemical modification with stabilizing agents, formation of cross-linked enzyme aggregates, pretreatment with polar organic solvents or enzymes combined with suitable surfactants to form microemulsions. The use of these enzyme preparations in ILs can dramatically increase the solvent tolerance, enhance activity as well as stability, and improve enantioselectivity. This perspective highlights a number of pronounced strategies being used successfully for activation and stabilization of enzymes in non-aqueous ILs media. This review is not intended to be comprehensive, but rather to present a general overview of the potential approaches to activate enzymes for diverse enzymatic processes and biotransformations in ILs.. |
| 338. |
Hiroki Abe, Masahiro GOTO, Noriho KAMIYA, Enzymatic single-step preparation of multifunctional proteins, Chem. Commun., 46, 7160-7162, 2010.05. |
| 339. |
Hidehiko Hirakawa, Noriho Kamiya, Yutaka Kawarabayasi, Teruyuki Nagamune, Artificial self-sufficient P450 in reversed micelles, Molecules, 10.3390/molecules15052935, 15, 5, 2935-2948, 2010.05, Cytochrome P450s are heme-containing monooxygenases that require electron transfer proteins for their catalytic activities. They prefer hydrophobic compounds as substrates and it is, therefore, desirable to perform their reactions in non-aqueous media. Reversed micelles can stably encapsulate proteins in nano-scaled water pools in organic solvents. However, in the reversed micellar system, when multiple proteins are involved in a reaction they can be separated into different micelles and it is then difficult to transfer electrons between proteins. We show here that an artificial self-sufficient cytochrome P450, which is an enzymatically crosslinked fusion protein composed of P450 and electron transfer proteins, showed micelle-size dependent catalytic activity in a reversed micellar system. Furthermore, the presence of thermostable alcohol dehydrogenase promoted the P450-catalyzed reaction due to cofactor regeneration.. |
| 340. |
Kyunga Sung, Noriho Kamiya, Noriyuki Kawata, Shinji Kamiya, Masahiro Goto, Functional glass surface displaying a glutamyl donor substrate for transglutaminase-mediated protein immobilization, Biotechnology Journal, 10.1002/biot.200900302, 5, 5, 456-462, 2010.05, A chemically modified glass surface displaying a glutamyl donor substrate peptide (Z-QG) was developed for microbial transglutaminase (MTG)-mediated immobilization of recombinant proteins tagged with an MTG-reactive lysine-containing substrate peptide (K-tag). To evaluate the surface modification conditions affecting the enzymatic protein immobilization, we employed an amino-modified 96-well glass plate as a base and prepared three types of glass surfaces displaying Z-QG. Validation of the Z-QG modified glass surfaces with recombinant enhanced green fluorescent proteins revealed that the insertion of a di(ethylene glycol) linker between the terminal Z-QG moiety and the base not only enhances enzymatic protein immobilization efficiency but also decreases nonselective protein adsorption. A bacterial alkaline phosphatase fused with a K-tag at the N terminus was also successfully immobilized to the designed glass surface, suggesting that the chemically modified glass surface displaying a spatially controlled glutamyl donor substrate is a potential platform for MTG-mediated fabrication of protein-based solid biomaterials.. |
| 341. |
Hidehiko Hirakawa, Noriho Kamiya, Yutaka Kawarabayasi, Teruyuki Nagamune, Artificial Self-Sufficient P450 in Reversed Micelles, MOLECULES, 10.3390/molecules15052935, 15, 5, 2935-2948, 2010.05, Cytochrome P450s are heme-containing monooxygenases that require electron transfer proteins for their catalytic activities. They prefer hydrophobic compounds as substrates and it is, therefore, desirable to perform their reactions in non-aqueous media. Reversed micelles can stably encapsulate proteins in nano-scaled water pools in organic solvents. However, in the reversed micellar system, when multiple proteins are involved in a reaction they can be separated into different micelles and it is then difficult to transfer electrons between proteins. We show here that an artificial self-sufficient cytochrome P450, which is an enzymatically crosslinked fusion protein composed of P450 and electron transfer proteins, showed micelle-size dependent catalytic activity in a reversed micellar system. Furthermore, the presence of thermostable alcohol dehydrogenase promoted the P450-catalyzed reaction due to cofactor regeneration.. |
| 342. |
Kyunga Sung, Noriho Kamiya, Noriyuki Kawata, Shinji Kamiya, Masahiro Goto, Functional glass surface displaying a glutamyl donor substrate for transglutaminase-mediated protein immobilization, BIOTECHNOLOGY JOURNAL, 10.1002/biot.200900302, 5, 5, 456-462, 2010.05, A chemically modified glass surface displaying a glutamyl donor substrate peptide (Z-QG) was developed for microbial transglutaminase (MTG)-mediated immobilization of recombinant proteins tagged with an MTG-reactive lysine-containing substrate peptide (K-tag). To evaluate the surface modification conditions affecting the enzymatic protein immobilization, we employed an amino-modified 96-well glass plate as a base and prepared three types of glass surfaces displaying Z-QG. Validation of the Z-QG modified glass surfaces with recombinant enhanced green fluorescent proteins revealed that the insertion of a di(ethylene glycol) linker between the terminal Z-QG moiety and the base not only enhances enzymatic protein immobilization efficiency but also decreases nonselective protein adsorption. A bacterial alkaline phosphatase fused with a K-tag at the N terminus was also successfully immobilized to the designed glass surface, suggesting that the chemically modified glass surface displaying a spatially controlled glutamyl donor substrate is a potential platform for MTG-mediated fabrication of protein-based solid biomaterials.. |
| 343. |
新規ダブルコーティング型キャリアの開発と非ウイルス性遺伝子デリバリーへの応用. |
| 344. |
独自のエマルション化技術を利用した新規ダブルコーティング型キャリアの開発. |
| 345. |
Fukiko Kubota, Yousuke Shimobori, Yusuke Koyanagi, Kojiro Shimojo, Noriho Kamiya, Masahiro Goto, Uphill transport of rare-earth metals through a highly stable supported liquid membrane based on an ionic liquid, analytical sciences, 10.2116/analsci.26.289, 26, 3, 289-290, 2010.03, We have developed a highly stable supported liquid membrane based on ionic liquids (ILs) for the separation of rare-earth metals, employing N,N-dioctyldiglycol amic acid as a mobile carrier. The quantitative transport of Y and Eu through the membrane was successfully attained, and separation from metal impurities, Zn, was efficiently accomplished. A membrane stable enough for long-term operation was constructible from imidazolium-based ILs having a longer alkyl chain, such as octyl or dodecyl groups in an imidazolium cation. 2010. |
| 346. |
Fukiko Kubota, Yousuke Shimobori, Yusuke Koyanagi, Kojiro Shimojo, Noriho Kamiya, Masahiro Goto, Uphill Transport of Rare-Earth Metals through a Highly Stable Supported Liquid Membrane Based on an Ionic Liquid, ANALYTICAL SCIENCES, 10.2116/analsci.26.289, 26, 3, 289-290, 2010.03, We have developed a highly stable supported liquid membrane based on ionic liquids (ILs) for the separation of rare-earth metals, employing N,N-dioctyldiglycol amic acid as a mobile carrier. The quantitative transport of Y and Eu through the membrane was successfully attained, and separation from metal impurities, Zn, was efficiently accomplished. A membrane stable enough for long-term operation was constructible from imidazolium-based ILs having a longer alkyl chain, such as octyl or dodecyl groups in an imidazolium cation.. |
| 347. |
Muhammad Moniruzzaman, Kazunori Nakashima, Noriho Kamiya, Masahiro Goto, Recent advances of enzymatic reactions in ionic liquids, Biochemical Engineering Journal, 10.1016/j.bej.2009.10.002, 48, 3, 295-314, 2010.02, The tremendous potential of room temperature ionic liquids as an alternative to environmentally harmful ordinary organic solvents is well recognized. Ionic liquids, having no measurable vapor pressure, are an interesting class of tunable and designer solvents, and they have been used extensively in a wide range of applications including enzymatic biotransformation. In fact, ionic liquids can be designed with different cation and anion combinations, which allow the possibility of tailoring reaction solvents with specific desired properties, and these unconventional solvent properties of ionic liquids provide the opportunity to carry out many important biocatalytic reactions that are impossible in traditional solvents. As compared to those observed in conventional organic solvents, the use of enzymes in ionic liquids has presented many advantages such as high conversion rates, high enantioselectivity, better enzyme stability, as well as better recoverability and recyclability. To date, a wide range of pronounced approaches have been taken to further improve the performance of enzymes in ionic liquids. This review presents the recent technological developments in which the advantages of ionic liquids as a medium for enzymes have been gradually realized.. |
| 348. |
Muhammad Moniruzzaman, Kazunori Nakashima, Noriho Kamiya, Masahiro Goto, Recent advances of enzymatic reactions in ionic liquids, BIOCHEMICAL ENGINEERING JOURNAL, 10.1016/j.bej.2009.10.002, 48, 3, 295-314, 2010.02, The tremendous potential of room temperature ionic liquids as an alternative to environmentally harmful ordinary organic solvents is well recognized. Ionic liquids, having no measurable vapor pressure, are an interesting class of tunable and designer solvents, and they have been used extensively in a wide range of applications including enzymatic biotransformation. In fact, ionic liquids can be designed with different cation and anion combinations, which allow the possibility of tailoring reaction solvents with specific desired properties, and these unconventional solvent properties of ionic liquids provide the opportunity to carry out many important biocatalytic reactions that are impossible in traditional solvents. As compared to those observed in conventional organic solvents, the use of enzymes in ionic liquids has presented many advantages such as high conversion rates, high enantioselectivity, better enzyme stability, as well as better recoverability and recyclability. To date, a wide range of pronounced approaches have been taken to further improve the performance of enzymes in ionic liquids. This review presents the recent technological developments in which the advantages of ionic liquids as a medium for enzymes have been gradually realized. (C) 2009 Elsevier B.V. All rights reserved.. |
| 349. |
新たなタンパク質複合化技術を用いた固定化酵素調製法. |
| 350. |
M. Moniruzzaman, Y. Tahara, M. Tamura, N. Kamiya, M. Goto, Ionic liquid-assisted transdermal delivery of sparingly soluble drugs, Chem. Commun., 46, 1452-1454, 2010.01. |
| 351. |
Teppei Niide, Hikaru Shiraki, Tatsuya Oshima, Yoshinari Baba, Noriho Kamiya, Masahiro Goto, Quaternary ammonium bacterial cellulose for adsorption of proteins, Solvent Extraction Research and Development, 10.15261/serdj.17.73, 17, 73-81, 2010.01, Bacterial cellulose (BC) has attracted attention for preparing advanced materials due to its microfibrous structure. In this study, quaternary ammonium bacterial cellulose (QABC) as well as quaternary ammonium plant cellulose (QAPC) as adsorbents for proteins have been prepared. Introduction of the quaternary ammonium group was conducted under different conditions to determine the best conditions for modification. The fibrous structures of QABC and QAPC were quite different and determined by the starting material. The adsorption capacities of hemoglobin and an anionic dye, thymol blue, on the adsorbents QABC and QAPC which were prepared under different conditions have been investigated. QABC showed a higher adsorption capacity for hemoglobin compared with QAPC, but a lower adsorption capacity for thymol blue.. |
| 352. |
Muhammad Moniruzzaman, Yoshiro Tahara, Miki Tamura, Noriho Kamiya, Masahiro Goto, Ionic liquid-assisted transdermal delivery of sparingly soluble drugs, Chemical Communications, 10.1039/b907462g, 46, 9, 1452-1454, 2010, We report the first successful application of a novel IL-assisted non-aqueous microemulsion stabilized by a blend of two nontoxic surfactants, polyoxyethylene sorbitan monooleate (Tween-80), and sorbitan laurate (Span-20) for transdermal delivery of acyclovir, which is insoluble or sparingly soluble in water and most common organic liquids.. |
| 353. |
Josui Shimada, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Design of a molecular detection system by a DNA-regulated enzyme conjugate, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 10.1016/j.jbiosc.2009.08.327, 108, S112-S112, 2009.11. |
| 354. |
Development and clarification of a novel transcutaneous protein delivery system and its mechanism by a solid-in-oil nanodispersion. |
| 355. |
Noriho Kamiya, Hiroki Abe, Masahiro Goto, Yukiko Tsuji, Hiroyuki Jikuya, New fluorescent substrates designed for covalent protein labeling catalyzed by microbial transglutaminase, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 10.1016/j.jbiosc.2009.08.283, 108, S97-S97, 2009.11. |
| 356. |
T. Mouri, T. Shimizu, N. Kamiya,* H. Ichinose, M. Goto*, Design of a cytochrome P450BM3 reaction system linked by two-step cofactor regeneration catalyzed by a soluble transhydrogenase and glycerol dehydrogenase, Biotechnol. Prog., 25, 1372-1378, 2009.10. |
| 357. |
Tsuyoshi Mouri, Takeshi Shimizu, Noriho Kamiya, Masahiro Goto, Hirofumi Ichinose, Design of a cytochrome P450BM3 reaction system linked by two-step cofactor regeneration catalyzed by a soluble transhydrogenase and glycerol dehydrogenase, Biotechnology Progress, 10.1002/btpr.231, 25, 5, 1372-1378, 2009.09, A cytochrome P450BM3-catalyzed reaction system linked by a two-step cofactor regeneration was investigated in a cell-free system. The two-step cofactor regeneration of redox cofactors, NADH and NADPH, was constructed by NAD+-dependent bacterial glycerol dehydrogenase (GLD) and bacterial soluble transhydrogenase (STH) both from Escherichia coli. In the present system, the reduced cofactor (NADH) was regenerated by GLD from the oxidized cofactor (NAD+) using glycerol as a sacrificial cosubstrate. The reducing equivalents were subsequently transferred to NADP+ by STH as a cycling catalyst. The resultant regenerated NADPH was used for the substrate oxidation catalyzed by cytochrome P450BM3. The initial rate of the P450BM3-catalyzed reaction linked by the two-step cofactor regeneration showed a slight increase (approximately twice) when increasing the GLD units 10-fold under initial reaction conditions. In contrast, a 10-fold increase in STH units resulted in about a 9-fold increase in the initial reaction rate, implying that transhydrogenation catalyzed by STH was the rate-determining step. In the system lacking the two-step cofactor regeneration, 34% conversion of 50 lM of a model substrate (p-nitrophenoxydecanoic acid) was attained using 50 μM NADPH. In contrast, with the two-step cofactor regeneration, the same amount of substrate was completely converted using 5 lM of oxidized cofactors (NAD+ and NADP+) within 1 h. Furthermore, a 10-fold dilution of the oxidized cofactors still led to approximately 20% conversion in 1 h. These results indicate the potential of the combination of GLD and STH for use in redox cofactor recycling with catalytic quantities of NAD+ and NADP +.. |
| 358. |
Fukiko Kubota, Yosuke Shimobori, Yusuke Koyanagi, Kazunori Nakashima, Kojiro Shimojo, Noriho Kamiya, Masahiro Goto, Extraction behavior of indium with TOPO into ionic liquids, Solvent Extraction Research and Development, 16, 151-155, 2009.09, The extraction behavior of In3+ into ionic liquids, [C nmim][Tf2N], was investigated with trioctylphosphine oxide (TOPO) as the extractant and compared with that of a conventional organic solvent system. In the n-dodecane system, the extractability of In3+ increased with increasing hydrochloric acid concentration in the feed aqueous phase. On the contrary when using ionic liquids, In3+ was more extracted at lower concentrations of hydrochloric acid. The contrast between the two systems is probably due to the difference in their extraction mechanisms. Extraction and separation properties of In3+ from Sn4+, Fe3+and Al3+ were also examined. Indium was quantitatively extracted into the ionic liquid phase and the separation efficiency from other metal ions, especially from Fe3+ was enhanced compared with that in the organic solvent system.. |
| 359. |
Noriho Kamiya, Hiroki Abe, Masahiro Goto, Yukiko Tsuji, Hiroyuki Jikuya, Fluorescent substrates for covalent protein labeling catalyzed by microbial transglutaminase, Organic and Biomolecular Chemistry, 10.1039/b904046c, 7, 17, 3407-3412, 2009.09, Novel small substrates with a variety of fluorophores were designed for the covalent labeling of proteins catalyzed by microbial transglutaminase (MTG). The new design is based on the flexibility in the substrate recognition of MTG for the substitution of the N-terminal protecting group of a conventional transglutaminase substrate, benzyloxycarbonyl-L-glutaminylglycine (Z-QG). Here we report for the first time that MTG can accept diverse fluorophores (dansyl, fluorescein, and rhodamine derivatives) in place of the benzyloxycarbonyl moiety when linked via a β-alanine or ε-aminocaproic acid linker. The utility of the new fluorescent substrates was demonstrated by site-specific, covalent and quantitative labeling of an MTG-reactive Lys-containing peptide tag fused to the N-terminus of a recombinant bacterial alkaline phosphatase with retention of target protein functionality.. |
| 360. |
Tsuyoshi Mouri, Takeshi Shimizu, Noriho Kamiya, Masahiro Goto, Hirofumi Ichinose, Design of a Cytochrome P450BM3 Reaction System Linked by Two-Step Cofactor Regeneration Catalyzed by a Soluble Transhydrogenase and Glycerol Dehydrogenase, BIOTECHNOLOGY PROGRESS, 10.1002/btpr.231, 25, 5, 1372-1378, 2009.09, A cytochrome P450BM3-catalyzed reaction system linked by a two-step cofactor regeneration was investigated in a cell-free system. The two-step cofactor regeneration of redox cofactors, NADH and NADPH, was constructed by NAD(+)-dependent bacterial glycerol dehydrogenase (GLD) and bacterial soluble transhydrogenase (STH) both from Escherichia coli. In the present system, the reduced cofactor (NADH) was regenerated by GLD from the oxidized cofactor (NAD(+)) using glycerol as a sacrificial cosubstrate. The reducing equivalents were subsequently transferred to NADP(+) by STH as a cycling catalyst. The resultant regenerated NADPH was used for the substrate oxidation catalyzed by cytochrome P450BM3. The initial rate of the P450BM3-catalyzed reaction linked by the two-step cofactor regeneration showed a slight increase (approximately twice) when increasing the GLD units 10-fold under initial reaction conditions. In contrast, a 10-fold increase in STH units resulted in about a 9-fold increase in the initial reaction rate, implying that transhydrogenation catalyzed by STH was the rate-determining step. In the system lacking the two-step cofactor regeneration, 34% conversion of 50 mu M of a model substrate (p-nitrophenoxydecanoic acid) was attained using 50 mu M NADPH. In contrast, with the two-step cofactor regeneration, the same amount of substrate was completely converted using 5 mu M of oxidized cofactors (NAD(+) and NADP(+)) within 1 h. Furthermore, a 10-fold dilution of the oxidized cofactors still led to approximately 20% conversion in 1 h. These results indicate the potential of the combination of GLD and STH for use in redox cofactor recycling with catalytic quantities of NAD(+) and NADP(+). (c) 2009 American Institute of Chemical Engineers Biotechnol. Prog., 25: 1372-1378, 2009. |
| 361. |
N. Kamiya,* Y. Shiotari, M. Tokunaga, H. Matsunaga, H. Yamanouchi, K. Nakano, M. Goto, Stimuli-responsive nanoparticles composed of naturally occurring amphiphilic proteins, Chem. Commun., 5287-5289, 2009.08. |
| 362. |
N. Kamiya,* H. Abe, M. Goto, Y. Tsuji, H. Jikuya, Fluorescent substrates for covalent protein labeling catalyzed by microbial transglutaminase, Org. Biomol. Chem., 7, 3407-3412, 2009.08. |
| 363. |
1Ep12 Development of a novel protein modification method by focusing tyrosine residue. |
| 364. |
K. Minamihata, N. Kamiya,* S. Kiyoyama, H. Sakuraba, T. Ohshima, M. Goto, Development of a novel immobilization method for enzymes from hyperthermophiles, Biotechnol. Lett., 31, 1037-1041, 2009.07. |
| 365. |
K. Nakashima, N. Kamiya, D. Koda, T. Maruyama, M. Goto, Enzyme encapsulation in microparticles composed of polymerized ionic liquids for highly active and reusable biocatalysts, Org. Biomol. Chem., 7, 2353-2358, 2009.07. |
| 366. |
Kosuke Minamihata, Masamichi Tokunaga, Noriho Kamiya, Shiro Kiyoyama, Haruhiko Sakuraba, Toshihisa Ohshima, Masahiro Goto, Development of a novel immobilization method for enzymes from hyperthermophiles, Biotechnology letters, 10.1007/s10529-009-9974-8, 31, 7, 1037-1041, 2009.07, Peptide tags containing tyrosines (Y-tag) were introduced at the C-terminus of a hyperthermophilic enzyme, alkaline phosphatase from Pyrococcus furiosus (PfuAP). Immobilization of the recombinant PfuAPs onto water-in-oil-in-water (W/O/W) type microcapsules was performed by an in situ polymerization method. All the recombinant PfuAPs prepared in this study were quantitatively immobilized onto microcapsules. The PfuAP-immobilized microcapsules showed no significant loss of enzymatic activity until the 5th round of assays. This result implies that the recombinant PfuAPs were covalently immobilized onto microcapsules. Immobilized PfuAP tagged with a Y-tag having the sequence GGYYY exhibited approximately a twofold higher catalytic activity compared with the wild-type PfuAP.. |
| 367. |
Kosuke Minamihata, Masamichi Tokunaga, Noriho Kamiya, Shiro Kiyoyama, Haruhiko Sakuraba, Toshihisa Ohshima, Masahiro Goto, Development of a novel immobilization method for enzymes from hyperthermophiles, BIOTECHNOLOGY LETTERS, 10.1007/s10529-009-9974-8, 31, 7, 1037-1041, 2009.07, Peptide tags containing tyrosines (Y-tag) were introduced at the C-terminus of a hyperthermophilic enzyme, alkaline phosphatase from Pyrococcus furiosus (PfuAP). Immobilization of the recombinant PfuAPs onto water-in-oil-in-water (W/O/W) type microcapsules was performed by an in situ polymerization method. All the recombinant PfuAPs prepared in this study were quantitatively immobilized onto microcapsules. The PfuAP-immobilized microcapsules showed no significant loss of enzymatic activity until the 5th round of assays. This result implies that the recombinant PfuAPs were covalently immobilized onto microcapsules. Immobilized PfuAP tagged with a Y-tag having the sequence GGYYY exhibited approximately a twofold higher catalytic activity compared with the wild-type PfuAP.. |
| 368. |
A transdermal delivery system of an ascorbic acid derivative utilizing solid-in-oil technique. |
| 369. |
Kazunori Nakashima, Noriho Kamiya, Daisuke Koda, Tatsuo Maruyama, Masahiro Goto, Enzyme encapsulation in microparticles composed of polymerized ionic liquids for highly active and reusable biocatalysts, Organic and Biomolecular Chemistry, 10.1039/b823064a, 7, 11, 2353-2358, 2009.06, Horseradish peroxidase (HRP) is encapsulated in polymerized ionic liquid microparticles (pIL-MP), which are prepared by polymerization of 1-vinyl-3-ethylimidazolium bromide in the presence of the crosslinker N,N′-methylenebis(acrylamide) in a concentrated water-in-oil (W/O) emulsion. pIL-MP encapsulating HRP chemically-modified with comb-shaped polyethylene glycol (PM13-HRP) exhibit excellent activity for guaiacol oxidation in an aqueous solution. The PM13-HRP in pIL-MP shows more than 2-fold higher activity than that of the enzyme encapsulated in a polyacrylamide microparticle. The catalytic activity declines with an increase in the crosslinker concentration of the pIL-MP, probably due to suppression of substrate diffusion. The activity of PM13-HRP in pIL-MP depends on the external environment of the gel (i.e. pH and temperature). The pIL-MP are easily recovered from the reaction mixture by centrifugation, which makes it possible to recycle the biocatalyst for repeated oxidation reactions.. |
| 370. |
S/O化技術の魅力と新しい経皮薬物送達システム実現の可能性. |
| 371. |
M.M.Zaman, N. Kamiya, M. Goto, Biocatalysis in Water-in-Ionic Liquid Microemulsions: A Case Study with Horseradish Peroxidase, Langmuir, 25, 977-982, 2009.04. |
| 372. |
タンパク質の経皮デリバリーの実現. |
| 373. |
M. Moniruzzaman, N. Kamiya, M. Goto, Biocatalysis in water-in-ionic liquid microemulsions
A case study with horseradish peroxidase, Langmuir, 10.1021/la803118q, 25, 2, 977-982, 2009.01, In this article we report the first results on the enzymatic activity of horseradish peroxidase (HRP) microencapsulated in water-in-ionic liquid (w/IL) microemulsions using pyrogallol as the substrate. Toward this goal, the system used in this study was composed of anionic surfactant AOT (sodium bis(2-ethyl-1-hexyl)sulfosuccinate)/hydrophobic IL [C8mim][Tf 2N] (1-octyl-3-methyl imidazolium bis(trifluoromethylsulfonyl)amide)/ water/l-hexanol. In this system, the catalytic activity of HRP was measured as a function of substrate concentrations, W0 (molar ratio of water to surfactant), pH, and 1-hexanol content. The curve of the activity-W0 profile was found to be hyperbolic for the new microemulsion. The apparent Michaelis-Menten kinetic parameters (Kcat and Km) were estimated and compared to those obtained from a conventional microemulsion. Apparently, it was found that HRP-catalyzed oxidation of pyrogallol by hydrogen peroxide in IL microemulsuions is much more effective than in a conventional AOT/water/isooctane microemulsion. The stability of HRP solubilized in the newly developed w/IL microemulsions was examined, and it was found that HRP retained almost 70% of its initial activity after incubation at 28 °C for 30 h.. |
| 374. |
M. Moniruzzaman, N. Kamiya, A. Goto, Biocatalysis in Water-in-Ionic Liquid Microemulsions: A Case Study with Horseradish Peroxidase, LANGMUIR, 10.1021/la803118q, 25, 2, 977-982, 2009.01, In this article we report die first results on the enzymatic activity of horseradish peroxidase (HRP) microencapsulated in water-in-ionic liquid (w/IL) microemulsions using pyrogallol as the substrate. Toward this goal, the system used in this study was composed of anionic surfactant AOT (sodium bis(2-ethyl-1-hexyl)sulfosuccinate)/hydrophobic IL [C(8)mim][Tf(2)N] (1-octyl-3-methyl imidazolium bis(trifluoromethylsulfonyl)amide)/water/1-hexanol. In this system, the catalytic activity of HRP was measured as a function of substrate concentrations, W(0) (molar ratio of water to surfactant), pH, and 1-hexanol content. The curve of the activity-W(0) profile was found to be hyperbolic for the new microemulsion. The apparent Michaelis-Menten kinetic parameters (k(cat) and K(m)) were estimated and compared to those obtained from a conventional microemulsion. Apparently, it-was found that HRP-catalyzed oxidation of pyrogallol by hydrogen peroxide in IL microemulsions is much more effective than in a conventional AOT/water/isooctane microemulsion. The stability of HRP solubilized in the newly developed W/IL microemulsions was examined, and it was found that HRP retained almost 70% of its initial activity after incubation at 28 degrees C for 30 h.. |
| 375. |
Noriho Kamiya, Yoshiaki Shiotari, Masamichi Tokunaga, Hideshi Matsunaga, Hirokazu Yamanouchi, Koji Nakano, Masahiro Goto, Stimuli-responsive nanoparticles composed of naturally occurring amphiphilic proteins, Chemical Communications, 10.1039/b909897f, 35, 5287-5289, 2009, Simple chemical cross-linking of β-casein micelles resulted in the formation of thermally responsive proteinaceous nanoparticles that exhibit lower critical solution temperatures.. |
| 376. |
Josui Shimada, Tatsuo Maruyama, Takuya Hosogi, Jo Tominaga, Noriho Kamiya, Masahiro Goto, Conjugation of DNA with protein using His-tag chemistry and its application to the aptamer-based detection system, Biotechnology letters, 10.1007/s10529-008-9784-4, 30, 11, 2001-2006, 2008.11, We propose a novel method to prepare a DNA-protein conjugate using histidine-tag (His-tag) chemistry. Oligo-DNA was modified with nitrilotriacetate (NTA), which has high affinity to a His-tag on recombinant protein via the complexation of Ni2+. Investigations using a microplate which displayed a complementary DNA-strand revealed that a NTA-modified DNA-protein conjugate was formed and immobilized in the presence of Ni2+ on the microplate. We then adopted alkaline phosphatase (AP) as a model protein, and application of the DNA-AP conjugate was demonstrated in a thrombin aptamer-based detection system with a detection limit of approximately 10 nM.. |
| 377. |
Josui Shimada, Tatsuo Maruyama, Takuya Hosogi, Jo Tominaga, Noriho Kamiya, Masahiro Goto, Conjugation of DNA with protein using His-tag chemistry and its application to the aptamer-based detection system, BIOTECHNOLOGY LETTERS, 10.1007/s10529-008-9784-4, 30, 11, 2001-2006, 2008.11, We propose a novel method to prepare a DNA-protein conjugate using histidine-tag (His-tag) chemistry. Oligo-DNA was modified with nitrilotriacetate (NTA), which has high affinity to a His-tag on recombinant protein via the complexation of Ni(2+). Investigations using a microplate which displayed a complementary DNA-strand revealed that a NTA-modified DNA-protein conjugate was formed and immobilized in the presence of Ni(2+) on the microplate. We then adopted alkaline phosphatase (AP) as a model protein, and application of the DNA-AP conjugate was demonstrated in a thrombin aptamer-based detection system with a detection limit of approximately 10 nM.. |
| 378. |
Yoshiro Tahara, Shota Honda, Noriho Kamiya, Hongyu Piao, Akihiko Hirata, Eiji Hayakawa, Takeru Fujii, Masahiro Goto, A solid-in-oil nanodispersion for transcutaneous protein delivery, Journal of Controlled Release, 10.1016/j.jconrel.2008.07.015, 131, 1, 14-18, 2008.10, Transcutaneous delivery attracts much attention but remains a challenging strategy for hydrophilic macromolecular drug administration. In the present study, we demonstrated that a solid-in-oil (S/O) nanodispersion, an oil-based nanodispersion of hydrophilic drugs, effectively enhanced the permeation of proteins into the skin. All of the different model proteins, FITC-labeled insulin (MW ca. 6 kDa), enhanced green fluorescent protein (EGFP, MW ca. 27 kDa) and horseradish peroxidase (HRP, MW ca. 40 kDa), permeated through the stratum corneum of Yucatan micropig skin in vitro by forming a S/O nanodispersion. The penetrated EGFP and HRP exhibited green fluorescence and catalytic activity, respectively, suggesting that these proteins can permeate into the skin in a functional form. The results indicated the potential utility of the S/O nanodispersion as a novel vehicle for transcutaneous protein delivery.. |
| 379. |
Yoshiro Tahara, Shota Honda, Noriho Kamiya, Hongyu Piao, Akihiko Hirata, Eiji Hayakawa, Takeru Fujii, Masahiro Goto, A solid-in-oil nanodispersion for transcutaneous protein delivery, JOURNAL OF CONTROLLED RELEASE, 10.1016/j.jconrel.2008.07.015, 131, 1, 14-18, 2008.10, Transcutaneous delivery attracts much attention but remains a challenging strategy for hydrophilic macromolecular drug administration. In the present study, we demonstrated that a solid-in-oil (S/O) nanodispersion, an oil-based nanodispersion of hydrophilic drugs, effectively enhanced the permeation of proteins into the skin. All of the different model proteins, FITC-labeled insulin (MW ca. 6 kDa), enhanced green fluorescent protein (EGFP, MW ca. 27 kDa) and horseradish peroxidase (HRP, MW ca. 40 kDa), permeated through the stratum corneum of Yucatan micropig skin in vitro by forming a S/O nanodispersion. The penetrated EGFP and HRP exhibited green fluorescence and catalytic activity, respectively, suggesting that these proteins can permeate into the skin in a functional form. The results indicated the potential utility of the S/O nanodispersion as a novel vehicle for transcutaneous protein delivery. (C) 2008 Elsevier B.V. All rights reserved.. |
| 380. |
Hiromu Yoshiura, Yoshiro Tahara, Masakazu Hashida, Noriho Kamiya, Akihiko Hirata, Takeru Fujii, Masahiro Goto, Design and in vivo evaluation of solid-in-oil suspension for oral delivery of human growth hormone, Biochemical Engineering Journal, 10.1016/j.bej.2008.04.001, 41, 2, 106-110, 2008.09, Solid-in-oil (S/O) suspension containing human growth hormone (hGH), in which hGH was complexed with an edible surfactant, was designed and validated for oral administration of hGH. To optimize the formulation procedures, protein release behavior from the S/O suspensions under physiological conditions was first investigated with horseradish peroxidase (HRP) as a model protein. Evaluation of functional integrity of HRP released from the HRP-surfactant complex suggested that the solubilization process partly impaired the specific activity of HRP, however, the addition of trehalose in the formulation process regained up to about 50% of the biological activity. On the basis of the data collected with HRP, a surfactant-hGH complex prepared under optimized conditions was suspended in soybean oil to formulate S/O suspensions. After oral administration of the S/O suspension to male New Zealand rabbits, we detected hGH in the serum with 3.3% bioavailability, suggesting that hGH can be orally delivered to the systemic circulation by the present formulation.. |
| 381. |
Fukiko Kubota, Yusuke Koyanagi, Kazunori Nakashima, Kojiro Shimojo, Noriho Kamiya, Masahiro Goto, Extraction of lanthanide ions with an organophosphorous extractant into ionic liquids, Solvent Extraction Research and Development, 15, 81-87, 2008.09, Extraction of lanthanide ions was carried out with the organophosphorous extractant PC-88A dissolved in an imidazolium-based ionic liquid. PC-88A was soluble in ionic liquids having long-chain alkyl substituents such as octyl and dodecyl groups, [C8mim] and [C12mim][TFSI], although short-chain imidazolium ionic liquids could not solubilize the extractant. The extraction equilibrium observed in the ionic liquid system was similar to that in the n-dodecane system, and the extraction behavior could be explained by the same proton exchange mechanism as for the conventional organic solvent system, although the extraction efficiency was not improved. An ionic liquid having shorter alkyl chain [C8mim] showed a higher extraction ability than [C12mim][TFSI]. Stripping of the metal ions from the ionic liquids was also examined. Quantitative stripping of the metal ions was achieved by nitric acid in the [C12mim][TFSI] system.. |
| 382. |
Hiromu Yoshiura, Yoshiro Tahara, Masakazu Hashida, Noriho Kamiya, Akihiko Hirata, Takeru Fujii, Masahiro Goto, Design and in vivo evaluation of solid-in-oil suspension for oral delivery of human growth hormone, BIOCHEMICAL ENGINEERING JOURNAL, 10.1016/j.bej.2008.04.001, 41, 2, 106-110, 2008.09, Solid-in-oil (S/O) suspension containing human growth hormone (hGH), in which hGH was complexed with an edible surfactant, was designed and validated for oral administration of hGH. To optimize the formulation procedures, protein release behavior from the S/O suspensions under physiological conditions was first investigated with horseradish peroxidase (HRP) as a model protein. Evaluation of functional integrity of HRP released from the HRP-surfactant complex suggested that the solubilization process partly impaired the specific activity of HRP, however, the addition of trehalose in the formulation process regained up to about 50% of the biological activity. On the basis of the data collected with HRP, a surfactant-hGH complex prepared under optimized conditions was suspended in soybean oil to formulate S/O suspensions. After oral administration of the S/O suspension to male New Zealand rabbits, we detected hGH in the serum with 3.3% bioavailability, suggesting that hGH can be orally delivered to the systemic circulation by the present formulation. (C) 2008 Elsevier B.V. All rights reserved.. |
| 383. |
Shiro Kiyoyama, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Microcapsulation of DNA and the adsorption of toxic substances, Journal of Microencapsulation, 10.1080/02652040801990741, 25, 5, 324-329, 2008.08, This study reports on deoxyribonucleic acid (DNA) immobilization in a microcapsule and the adsorption of toxic substances. Salmon testes DNA was immobilized using a microencapsulation method and the immobilization efficiency of DNA, stability of microcapsulated DNA and adsorption properties of toxic substances were investigated under various experimental conditions. This study succeeded in immobilizing over 90% of DNA in the DNA dosage based on the thymine base and the microcapsulation of DNA showed high reproducibility for immobilization efficiency. Immobilized DNA in the microcapsules did not leak, regardless of the salt concentration and pH values in the external aqueous phase. Microcapsulated DNA could selectively adsorb toxic substances that have a planar structure. Based on these results, it was shown that the microcapsulated DNA is a good adsorbent for toxic substances and the microcapsulation method is useful for immobilizing DNA in a polymer matrix.. |
| 384. |
H. Piao, N. Kamiya, A. Hirata, T. Fujii, I. Shimizu, S. Ito, M. Goto, Reduction of gastric ulcerogenicity during multiple administration of diclofenac sodium by a novel solid-in-oil suspension., Pharm. Devel. Technol., vol.12, 321-325 (2007), 2008.07. |
| 385. |
Yusuke Tanaka, Satoshi Doi, Noriho Kamiya, Noriyuki Kawata, Shinji Kamiya, Kenichi Nakama, Masahiro Goto, A chemically modified glass surface that facilitates transglutaminase- mediated protein immobilization, Biotechnology letters, 10.1007/s10529-008-9656-y, 30, 6, 1025-1029, 2008.06, An amino-modified glass surface for enzymatic protein immobilization by microbial transglutaminase (MTG) was developed. Diamine substrates with secondary amino groups in the linker moiety, like triethylenetetramine (TETA), exhibited at most a 2-fold higher reactivity in the MTG-catalyzed reaction compared to those with the alkyl linker. A 96-well glass plate was subsequently modified with selected diamine substrates. Validation of the modified surface by enzymatic immobilization of enhanced green fluorescent protein tagged with a glutamine donor-substrate peptide (LLQG) of MTG revealed that the protein loading onto the TETA-modified glass surface was approximately 15-fold higher than that on the unmodified one.. |
| 386. |
Noriho Kamiya, Yuichi Matsushita, Misa Hanaki, Kazunori Nakashima, Mamiko Narita, Masahiro Goto, Haruo Takahashi, Enzymatic in situ saccharification of cellulose in aqueous-ionic liquid media, Biotechnology letters, 10.1007/s10529-008-9638-0, 30, 6, 1037-1040, 2008.06, The enzymatic saccharification of a model cellulosic substrate, Avicel PH-101, using an ionic liquid (IL), 1-ethyl-3-methylimidazolium diethylphosphate, was explored. After mixing the IL solution of cellulose with different volumes of 10 mM citrate buffer (pH 5.0), cellulase was directly added to the aqueous-IL mixture at 40°C. When the volume of IL to water was greater than 3:2, little cellulase activity was observed. However, decreasing the volume ratio markedly enhanced enzymatic activity: an IL to water ratio of 1:4 (v/v) resulted in over 70% of the starting amount of cellulose (10 mg/ml) being converted to glucose and cellobiose.. |
| 387. |
Yusuke Tanaka, Satoshi Doi, Noriho Kamiya, Noriyuki Kawata, Shinji Kamiya, Kenichi Nakama, Masahiro Goto, A chemically modified glass surface that facilitates transglutaminase-mediated protein immobilization, BIOTECHNOLOGY LETTERS, 10.1007/s10529-008-9656-y, 30, 6, 1025-1029, 2008.06, An amino-modified glass surface for enzymatic protein immobilization by microbial transglutaminase (MTG) was developed. Diamine substrates with secondary amino groups in the linker moiety, like triethylenetetramine (TETA), exhibited at most a 2-fold higher reactivity in the MTG-catalyzed reaction compared to those with the alkyl linker. A 96-well glass plate was subsequently modified with selected diamine substrates. Validation of the modified surface by enzymatic immobilization of enhanced green fluorescent protein tagged with a glutamine donor-substrate peptide (LLQG) of MTG revealed that the protein loading onto the TETA-modified glass surface was approximately 15-fold higher than that on the unmodified one.. |
| 388. |
Noriho Kamiya, Yuichi Matsushita, Misa Hanaki, Kazunori Nakashima, Mamiko Narita, Masahiro Goto, Haruo Takahashi, Enzymatic in situ saccharification of cellulose in aqueous-ionic liquid media, BIOTECHNOLOGY LETTERS, 10.1007/s10529-008-9638-0, 30, 6, 1037-1040, 2008.06, The enzymatic saccharification of a model cellulosic substrate, Avicel PH- 101, using an ionic liquid ( IL), 1- ethyl- 3- methylimidazolium diethylphosphate, was explored. After mixing the IL solution of cellulose with different volumes of 10 mM citrate buffer ( pH 5.0), cellulase was directly added to the aqueous- IL mixture at 40 degrees C. When the volume of IL to water was greater than 3: 2, little cellulase activity was observed. However, decreasing the volume ratio markedly enhanced enzymatic activity: an IL to water ratio of 1: 4 ( v/ v) resulted in over 70% of the starting amount of cellulose ( 10 mg/ ml) being converted to glucose and cellobiose.. |
| 389. |
Muhammad Moniruzzaman, Noriho Kamiya, Kazunori Nakashima, Masahiro Goto, Water-in-ionic liquid microemulsions as a new medium for enzymatic reactions, Green Chemistry, 10.1039/b802501k, 10, 5, 497-450, 2008.05, The insolubility of enzymes in most ionic liquids has been overcome by the formation of aqueous microemulsion droplets in a hydrophobic ionic liquid stabilized by a layer of anionic surfactant sodium bis(2-ethyl-1-hexyl) sulfosuccinate (AOT) in the presence of 1-hexanol as a cosurfactant and the catalytic activity of one of the enzymes studied (lipase PS) became higher than in microemulsions of AOT in isooctane.. |
| 390. |
Hongyu Piao, Noriho Kamiya, Akihiko Hirata, Takeru Fujii, Masahiro Goto, A novel solid-in-oil nanosuspension for transdermal delivery of diclofenac sodium, Pharmaceutical Research, 10.1007/s11095-007-9445-7, 25, 4, 896-901, 2008.04, Purpose. We formulated a solid-in-oil nanosuspension (SONS) as a novel transdermal delivery carrier for diclofenac sodium (DFNa). The basic transdermal characteristics of the SONS were evaluated using a Yucatan micropig (YMP) skin model. Methods. DFNa-sucrose erucate (i.e. surfactant) complexes were prepared via the formation of a water-in-oil emulsion. The complexes were suspended in isopropyl myristate (IPM) to form a SONS. The basic transdermal characteristics of the SONS were examined using full-thickness YMP dorsal skin in a Franz-type diffusion cell. DFNa powder suspended in IPM without complex formation was used as a control. The effect of the weight ratio of surfactant to DFNa on DFNa penetration of the skin was evaluated. Results. DFNa was successfully dispersed into IPM as a nanosized suspension via complex formation with sucrose erucate. The resultant SONS increased the permeability flux of DFNa across the YMP skin by up to 3.8-fold compared with the control. The size of the SONS depended on the weight ratio of the surfactant to DFNa. The optimal weight ratio for the highest DFNa permeation was 8.8, at which point the mean diameter of the SONS was 14.4 nm. Conclusion. The SONS formulation can enhance the percutaneous absorption of DFNa.. |
| 391. |
Muhammad Moniruzzaman, Noriho Kamiya, Kazunori Nakashima, Masahiro Goto, Formation of reverse micelles in a room-temperature ionic liquid, ChemPhysChem, 10.1002/cphc.200700802, 9, 5, 689-692, 2008.04, (Graph Presented) Nanopools of water in ionic liquids: The surfactant AOT forms reverse micelles with water domains in a hydrophobic ionic liquid in the presence of 1-hexanol (see optically transparent sample B, in figure). These nanopools increase in size with increasing water content.. |
| 392. |
Hongyu Piao, Noriho Kamiya, Akihiko Hirata, Takeru Fujii, Masahiro Goto, A novel solid-in-oil nanosuspension for transdermal delivery of diclofenac sodium, PHARMACEUTICAL RESEARCH, 10.1007/s11095-007-9445-7, 25, 4, 896-901, 2008.04, Purpose. We formulated a solid-in-oil nanosuspension (SONS) as a novel transdermal delivery carrier for diclofenac sodium (DFNa). The basic transdermal characteristics of the SONS were evaluated using a Yucatan micropig (YMP) skin model.
Methods. DFNa-sucrose erucate (i.e. surfactant) complexes were prepared via the formation of a water-in-oil emulsion. The complexes were suspended in isopropyl myristate (IPM) to form a SONS. The basic transdermal characteristics of the SONS were examined using full-thickness YMP dorsal skin in a Franz-type diffusion cell. DFNa powder suspended in IPM without complex formation was used as a control. The effect of the weight ratio of surfactant to DFNa on DFNa penetration of the skin was evaluated.
Results. DFNa was successfully dispersed into IPM as a nanosized suspension via complex formation with sucrose erucate. The resultant SONS increased the permeability flux of DFNa across the YMP skin by up to 3.8-fold compared with the control. The size of the SONS depended on the weight ratio of the surfactant to DFNa. The optimal weight ratio for the highest DFNa permeation was 8.8, at which point the mean diameter of the SONS was 14.4 nm.
Conclusion. The SONS formulation can enhance the percutaneous absorption of DFNa.. |
| 393. |
Muhammad Moniruzzaman, Kamiya Noriho, Kazunori Nakashima, Masahiro Goto, Formation of reverse micelles in a room-temperature ionic liquid, CHEMPHYSCHEM, 10.1002/cphc.200700802, 9, 5, 689-692, 2008.04, (Graph Presented) Nanopools of water in ionic liquids: The surfactant AOT forms reverse micelles with water domains in a hydrophobic ionic liquid in the presence of 1-hexanol (see optically transparent sample B, in figure). These nanopools increase in size with increasing water content. © 2008 Wiley-VCH Verlag GmbH & Co. KGaA.. |
| 394. |
Shiro Kiyoyama, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Immobilization of proteins into microcapsules and their adsorption properties with respect to precious-metal ions, Industrial and Engineering Chemistry Research, 10.1021/ie0712636, 47, 5, 1527-1532, 2008.03, We report on protein immobilization in a microcapsule and its adsorption performance with respect to preciousmetal ions. Proteins including lysozyme, bovine serum albumin, chicken egg albumin, and soybean protein were immobilized by a microencapsulation method, and their adsorption properties with respect to ions of both precious metals (gold, platinum, and palladium) and base metals (zinc and copper) were investigated under various preparation conditions. Immobilized proteins could selectively adsorb precious-metal ions over base-metal ions. High immobilization efficiencies were obtained using lysozyme and soybean protein. The immobilization efficiency of the proteins was controlled by the surfactant concentration in the organic phase, the protein concentration, and the outer-aqueous-phase composition. We succeeded in immobilizing over 90% of the protein in the protein dose by optimizing the preparation conditions. Furthermore, the preciousmetal ions adsorbed by the immobilized proteins were completely recovered with thiourea. No leakage of proteins was observed after the adsorption and desorption experiments. These results indicate that the microcapsules immobilizing proteins prepared in the present study can be used as reusable precious-metal-selective absorbents.. |
| 395. |
Kazunori Nakashima, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Spectrophotometric assay for protease activity in ionic liquids using chromogenic substrates, Analytical Biochemistry, 10.1016/j.ab.2007.11.015, 374, 2, 285-290, 2008.03, A new spectrophotometric assay has been developed to evaluate protease activity in ionic liquids (ILs). The assay consists of two strategies to enable real-time spectrometric analysis of enzymatic reaction in ILs. First, enzymes are modified with a comb-shaped poly(ethylene glycol), PM13, to obtain a transparent enzyme solution in IL. Second, a chromogenic substrate is used to follow the enzymatic reaction in IL. p-Nitroaniline-derivatized substrates are subjected to protease-catalyzed alcoholysis to release chromogenic p-nitroaniline that can be quantitatively detected by a UV-Vis spectrophotometer. By using this method, we can evaluate protease activity in ILs quite easily without separation of products from the reaction mixture. The availability of the novel assay system was demonstrated in a kinetic analysis of subtilisin-catalyzed reaction in the IL 1-ethyl-3-methylimidazolium bis(trifluoromethanesulfonyl)imide ([Emim][Tf2N]) under different reaction conditions. Because two different serine proteases, subtilisin and α-chymotrypsin, substantially retained its original substrate specificity in the IL, the assay can be extended to other enzymes by using suitable chromogenic substrates.. |
| 396. |
Kazunori Nakashima, Tatsuo Maruyama, Masahiro Goto, Noriho Kamiya, A rapid assay of protease activity in ionic liquids using chromogenic substrates, Analytical Biochemistry, Vol. 374 Issue: 2 Pages: 2, 2008.03. |
| 397. |
Shiro Kiyoyama, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Immobilization of proteins into microcapsules and their adsorption properties with respect to precious-metal ions, INDUSTRIAL & ENGINEERING CHEMISTRY RESEARCH, 10.1021/ie0712636, 47, 5, 1527-1532, 2008.03, We report on protein immobilization in a microcapsule and its adsorption performance with respect to precious-metal ions. Proteins including lysozyme, bovine serum albumin, chicken egg albumin, and soybean protein were immobilized by a microencapsulation method, and their adsorption properties with respect to ions of both precious metals (gold, platinum, and palladium) and base metals (zinc and copper) were investigated under various preparation conditions. Immobilized proteins could selectively adsorb precious-metal ions over base-metal ions. High immobilization efficiencies were obtained using lysozyme and soybean protein. The immobilization efficiency of the proteins was controlled by the surfactant concentration in the organic phase, the protein concentration, and the outer-aqueous-phase composition. We succeeded in immobilizing over 90% of the protein in the protein dose by optimizing the preparation conditions. Furthermore, the precious-metal ions adsorbed by the immobilized proteins were completely recovered with thiourea. No leakage of proteins was observed after the adsorption and desorption experiments. These results indicate that the microcapsules immobilizing proteins prepared in the present study can be used as reusable precious-metal-selective absorbents.. |
| 398. |
Kazunorl Nakashima, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Spectrophotometric assay for protease activity in ionic liquids using chromogenic substrates, ANALYTICAL BIOCHEMISTRY, 10.1016/j.ab.2007.11.015, 374, 2, 285-290, 2008.03, A new spectrophotometric assay has been developed to evaluate protease activity in ionic liquids (ILs). The assay consists of two strategies to enable real-time spectrometric analysis of enzymatic reaction in ILs. First, enzymes are modified with a comb-shaped poly(ethylene glycol), PM13, to obtain a transparent enzyme solution in IL. Second, a chromogenic substrate is used to follow the enzymatic reaction in IL. p-Nitroaniline-derivatized substrates are subjected to protease-catalyzed alcoholysis to release chromogenic p-nitroaniline that can be quantitatively detected by a UV-Vis spectrophotometer. By using this method, we can evaluate protease activity in ILs quite easily without separation of products from the reaction mixture. The availability of the novel assay system was demonstrated in a kinetic analysis of subtilisin-catalyzed reaction in the IL 1-ethyl-3-methylimidazolium bis(trifluoromethanesulfonyl)imide ([Emim][Tf2N]) under different reaction conditions. Because two different serine proteases, subtilisin and alpha-chymotrypsin, substantially retained its original substrate specificity in the IL, the assay can be extended to other enzymes by using suitable chromogenic substrates. (C) 2007 Elsevier Inc. All rights reserved.. |
| 399. |
N. Kamiya, S. Doi, Y. Tanaka, H. Ichinose, M. Goto, Functional immobilization of recombinant alkaline phosphatases bearing a glutamyl donor substrate peptide of microbial transglutaminase, J. Biosci. Bioeng., 104, 195-199, 2007.09. |
| 400. |
Noriho Kamiya, Satoshi Doi, Yusuke Tanaka, Hirofumi Ichinose, Masahiro Goto, Functional immobilization of recombinant alkaline phosphatases bearing a glutamyl donor substrate peptide of microbial transglutaminase, Journal of Bioscience and Bioengineering, 10.1263/jbb.104.195, 104, 3, 195-199, 2007.09, Covalent and site-specific protein immobilization catalyzed by microbial transglutaminase (MTG) was investigated using recombinant Escherichia coli alkaline phosphatase (AP) tagged with a glutamyl donor substrate peptide (MLAQGS) of MTG. A polystyrene surface physically coated with β-casein or bovine serum albumin (BSA) was employed as an MTG-specific surface displaying reactive lysine residues. MTG-mediated protein immobilization through catalytic ε-(γ-glutamyl)lysine bond formation between the peptide tag of recombinant APs and β-casein- or BSA-coated surface was verified by the detection of AP activity on the surface. It was found that the length and the insertion position of the peptide tag did not significantly affect the efficacy of enzymatic immobilization of the recombinant APs. On the other hand, pH and ionic strength in the reaction media had crucial effects on the immobilization yields. Interestingly, the optimum pH range of MTG-mediated protein immobilization differed markedly from that for an MTG-catalyzed reaction in aqueous solution. The results suggest that the concentration of reactive species due to electrostatic interaction between the enzyme-substrate intermediate and the protein-adsorbed surface is a key factor governing MTG catalysis at a solid surface.. |
| 401. |
Hidehiko Hirakawa, Noriho Kamiya, Tsutomu Tanaka, Teruyuki Nagamune, Intramolecular electron transfer in a cytochrome P450cam system with a site-specific branched structure, Protein Engineering, Design and Selection, 10.1093/protein/gzm045, 20, 9, 453-459, 2007.09, Cytochrome P450 (P450) is an attractive oxygenase due to the diverse catalytic reactions and the broad substrate specificity. Class I P450s require an excess concentration (more than 10 times) of iron-sulfur proteins, which transfer electrons to P450s, to attain the maximum catalytic activity and this requirement is a critical bottleneck for practical applications. Here, we show a site-specific branched fusion protein of P450 with its electron transfer proteins using enzymatic cross-linking with transglutaminase. A branched fusion protein of P450 from Pseudomonas putida (P450cam), which was composed of one molecule each of P450cam, putidaredoxin (Pdx) and Pdx reductase, showed higher catalytic activity (306 min-1) and coupling efficiency (99%) than the equimolar reconstitution system due to the intramolecular electron transfer. The unique site-specific branched structure simply increased local concentration of proteins without denaturation of each protein. Therefore, enzymatic post-translational protein manipulation can be a powerful alternative to conventional strategies for the creation of multicomponent enzyme systems with novel proteinaceous architecture.. |
| 402. |
Noriho Kamiya, Satoshi Doi, Yusuke Tanaka, Hirofumi Ichinose, Masahiro Goto, Functional immobilization of recombinant alkaline phosphatases bearing a glutarnyl donor substrate peptide of microbial transglutaminase, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 10.1263/jbb.104.195, 104, 3, 195-199, 2007.09, Covalent and site-specific protein immobilization catalyzed by microbial transglutaminase (MTG) was investigated using recombinant Escherichia coli alkaline phosphatase (AP) tagged with a glutamyl donor substrate peptide (MLAQGS) of MTC. A polystyrene surface physically coated with beta-casein or bovine serum albumin (BSA) was employed as an MTG-specific surface displaying reactive lysine residues. MTG-mediated protein immobilization. through catalytic epsilon-(gamma-glutamyl)lysine bond formation between the peptide tag of recombinant APs and beta-cascin- or BSA-coated surface was verified by the detection of AP activity on the surface. It was found that the length and the insertion position of the peptide tag did not significantly affect the efficacy of enzymatic immobilization of the recombinant APs. On the other hand, pH and ionic strength in the reaction media had crucial effects on the immobilization yields. Interestingly, the optimum pH range of MTG-mediated protein immobilization differed markedly from that for an MTG-catalyzed reaction in aqueous solution. The results suggest that the concentration of reactive species due to electrostatic interaction between the enzyme-substrate intermediate and the protein-adsorbed surface is a key factor governing MTG catalysis at a solid surface.. |
| 403. |
Hidehiko Hirakawa, Noriho Kamiya, Tsutomu Tanaka, Teruyuki Nagamune, Intramolecular electron transfer in a cytochrome P450cam system with a site-specific branched structure, PROTEIN ENGINEERING DESIGN & SELECTION, 10.1039/protein/gzm045, 20, 9, 453-459, 2007.09, Cytochrome P450 (P450) is an attractive oxygenase due to the diverse catalytic reactions and the broad substrate specificity. Class I P450s require an excess concentration (more than 10 times) of iron-sulfur proteins, which transfer electrons to P450s, to attain the maximum catalytic activity and this requirement is a critical bottleneck for practical applications. Here, we show a site-specific branched fusion protein of P450 with its electron transfer proteins using enzymatic cross-linking with transglutaminase. A branched fusion protein of P450 from Pseudomonas putida (P450cam), which was composed of one molecule each of P450cam, putidaredoxin (Pdx) and Pdx reductase, showed higher catalytic activity (306 min(-1)) and coupling efficiency (99%) than the equimolar reconstitution system due to the intramolecular electron transfer. The unique site-specific branched structure simply increased local concentration of proteins without denaturation of each protein. Therefore, enzymatic post-translational protein manipulation can be a powerful alternative to conventional strategies for the creation of multicomponent enzyme systems with novel proteinaceous architecture.. |
| 404. |
Fukiko Kubota, Yusuke Koyanagi, Kazunori Nakashima, Kojiro Shimojo, Noriho Kamiya, Masahiro Goto, Extraction of cytochrome c by a functionalized ionic liquid containing a crown ether, Solvent Extraction Research and Development, 14, 115-120, 2007.08, To verify the applicability of functionalized ionic liquids, so-called 'task-specific ionic liquids', to liquid-liquid extraction as solvents possessing extraction ability, we carried out the extraction of a protein cytochrome c (Cyt-c) with an ionic liquid with an attached crown ether moiety in the imidazolium cation. The functionalized ionic liquid, [18C6mim][PF 6], was dissolved in two different ionic liquids as diluents, one having a hydroxyethyl group, [C2OHmim][Tf2N], and the other an ethyl group, [C2mim] [Tf2N], respectively. The transfer of Cyt-c from the aqueous phase to the ionic liquid phase took place with [18C6mim][PF6] in the [C2OHmim][Tf2N] system. The functionalized ionic liquid with the crown ether attached provided the similar extraction behavior to that using dicyclohexano-18-crown-6 (DCH18C6) as the extradant. Although the extraction ability was lower than that of DCH18C6, the functionalized ionic liquid system gave a better stripping performance than the ordinary crown ether system. The extraction efficiency was not influenced by operating temperature when the concentration of [18C6mim] was high whereas it was greatly affected by temperature at low [18C6mim] concentration. The functionalized ionic liquids showed an extremely higher extraction performance compared to that of the ordinary ionic liquids, which have no coordination sites for the target molecule.. |
| 405. |
H. Yoshiura, M. Hashida, N. Kamiya, M. Goto, Factors affecting protein release behavior from surfactant-protein complexes under physiological conditions., Int. J. Pharm., 338, 174-179 (2007), 2007.07. |
| 406. |
Hiromu Yoshiura, Masakazu Hashida, Noriho Kamiya, Masahiro Goto, Factors affecting protein release behavior from surfactant-protein complexes under physiological conditions, International Journal of Pharmaceutics, 10.1016/j.ijpharm.2007.01.041, 338, 1-2, 174-179, 2007.06, Protein release behavior from its complex with edible surfactants was investigated under physiological conditions using hen egg lysozyme and Aspergillus niger glucose oxidase as model proteins. It revealed that protein release rates could be controlled by hydrophobicity of surfactants and the molar ratio of proteins to surfactants in the preparation of the complexes. Evaluation of functional integrity of a protein on the basis of specific activity of an enzyme released from the complex suggested that lower hydrophobicity of surfactants led to higher retention of catalytic activity. In addition, it was found that protein release rates from the complexes were correlated with the aqueous droplet size of water-in-oil emulsions in the preparation of the complexes. The results suggest the potential of surfactant-protein complexes in pharmaceutical formulations for mucosal delivery of therapeutic proteins.. |
| 407. |
Hiromu Yoshiura, Masakazu Hashida, Noriho Kamiya, Masahiro Goto, Factors affecting protein release behavior from surfactant-protein complexes under physiological conditions, INTERNATIONAL JOURNAL OF PHARMACEUTICS, 10.1016/j.ijpharm.2007.01.041, 338, 1-2, 174-179, 2007.06, Protein release behavior from its complex with edible surfactants was investigated under physiological conditions using hen egg lysozyme and Aspergillus niger glucose oxidase as model proteins. It revealed that protein release rates could be controlled by hydrophobicity of surfactants and the molar ratio of proteins to surfactants in the preparation of the complexes. Evaluation of functional integrity of a protein on the basis of specific activity of an enzyme released from the complex suggested that lower hydrophobicity of surfactants led to higher retention of catalytic activity. In addition, it was found that protein release rates from the complexes were correlated with the aqueous droplet size of water-in-oil emulsions in the preparation of the complexes. The results suggest the potential of surfactant-protein complexes in pharmaceutical formulations for mucosal delivery of therapeutic proteins. (c) 2007 Elsevier B.V. All rights reserved.. |
| 408. |
Y. Tanaka, Y. Tsuruda, M. Nishi, N. Kamiya, M. Goto, Exploring enzymatic catalysis at a solid surface: a case study with transglutaminase-mediated protein immobilization, Org. Biomol. Chem., 5, 1764-1770, 2007.05. |
| 409. |
Jo Tominaga, Noriho Kamiya, Masahiro Goto, An enzyme-labeled protein polymer bearing pendent haptens, Bioconjugate Chemistry, 10.1021/bc060161d, 18, 3, 860-865, 2007.05, A new methodology for the preparation of enzyme-labeled protein polymers bearing pendent haptens was developed through the combination of chemical modification and posttranslational protein modification catalyzed by microbial transglutaminase (MTG). As a model hapten, trinitrobenzene (TNB) was chosen and chemically conjugated with the accessible Lys residues of β-casein. The resultant trinitrophenylated β-casein was further modified with formaldehyde to render the residual Lys residues inert toward self-cross-linking by MTG. Escherichia coli alkaline phosphatase (AP), comprising a specific peptide tag carrying a MTG-reactive Lys residue, was then conjugated to the Gln residues in β-casein-TNB conjugates. The resultant AP-labeled β-casein-bearing pendent TNB moieties (AP-βCT) showed comparable specific activity with native AP. It was found that only the AP-βCT with a sufficient number of pendent TNBs are capable of binding to a surface adsorbed with anti-TNP and anti-TNT antibodies, indicating the presence of polyvalent interactions. The utility of AP-βCT was demonstrated by competitive immunoassays for trinitrophenol (TNP) and trinitrotoluene (TNT), with detection limits of 0.99 μg/L and 0.18 μg/L, respectively. The present study demonstrates the potential of dual labeling of protein scaffolds by chemical and enzymatic protein manipulation to create a new proteinaceous architecture.. |
| 410. |
Hongyu Piao, Akihiko Hirata, Hideakira Yokoyama, Takeru Fujii, Ichiro Shimizu, Susumu Ito, Noriho Kamiya, Masahiro Goto, Reduction of gastric ulcerogenicity during multiple administration of diclofenac sodium by a novel solid-in-oil suspension, Pharmaceutical Development and Technology, 10.1080/10837450701247517, 12, 3, 321-325, 2007.05, This article reports a significant reduction of gastric ulcerogenicity by complex formation of a nonsteroidal anti-inflammatory drug with surfactants. Diclofenac sodium (DFNa) was suspended in medium chain triglyceride (MCT) by forming a complex with an edible lipophilic surfactant. Two types of suspensions, prepared through a membrane emulsification with different pore sizes, were evaluated according to the degree of gastric damage following multiple oral administration in rats. It was shown that gastric ulcerogenicity of DFNa was reduced by the surfactant-drug complexes, at doses up to 12 mg/kg, whereas severe gastric damage was observed upon oral administration of the aqueous solution at doses of 6 mg/kg. Comparable blood levels of DFNa were observed after administration of solution and suspension formulations.. |
| 411. |
Jo Tominaga, Noriho Kamiya, Masahiro Goto, An enzyme-labeled protein polymer bearing pendent haptens, BIOCONJUGATE CHEMISTRY, 10.1021/bc060161d, 18, 3, 860-865, 2007.05, A new methodology for the preparation of enzyme-labeled protein polymers bearing pendent haptens was developed through the combination of chemical modification and posttranslational protein modification catalyzed by microbial transglutaminase (MTG). As a model hapten, trinitrobenzene (TNB) was chosen and chemically conjugated with the accessible Lys residues of beta-casein. The resultant trinitrophenylated beta-casein was further modified with formaldehyde to render the residual Lys residues inert toward self-cross-linking by MTG. Escherichia coli alkaline phosphatase (AP), comprising a specific peptide tag carrying a MTG-reactive Lys residue, was then conjugated to the Gln residues in beta-casein-TNB conjugates. The resultant AP-labeled beta-casein-bearing pendent TNB moieties (AP-beta CT) showed comparable specific activity with native AP. It was found that only the AP-beta CT with a sufficient number of pendent TNBs are capable of binding to a surface adsorbed with anti-TNP and anti-TNT antibodies, indicating the presence of polyvalent interactions. The utility of AP-beta CT was demonstrated by competitive immunoassays for trinitrophenol (TNP) and trinitrotoluene (TNT), with detection limits of 0.99 mu g/L and 0.18 mu g/L, respectively. The present study demonstrates the potential of dual labeling of protein scaffolds by chemical and enzymatic protein manipulation to create a new proteinaceous architecture.. |
| 412. |
J. Tominaga, N. Kamiya, M. Goto, An enzyme-labeled protein polymer bearing pendant haptens, Bioconjugate Chem., 18, 860-865, 2007.03. |
| 413. |
T. Maruyama, Y. Shimada, H. Matsushita, N. Kamiya, M. Goto, Proteins and protein-rich biomass as environmental-friendly adsorbents selective for precious metal ions, Env. Sci. Technol., 41, 1359-1364, 2007.02. |
| 414. |
Tatsuo Maruyama, Hironari Matsushita, Yukiko Shimada, Ichiro Kamata, Misa Hanaki, Saori Sonokawa, Noriho Kamiya, Masahiro Goto, Proteins and protein-rich biomass as environmentally friendly adsorbents selective for precious metal ions, Environmental Science and Technology, 10.1021/es061664x, 41, 4, 1359-1364, 2007.02, Proteins exhibit specific interactions with various metal ions, which play important roles in a living cell. Here, we found that various proteins selectively adsorbed precious metal ions at a wide range of pH values. Studies on protein sequences and on synthesized peptides revealed that a histidine-containing sequence had specific interactions with precious metal ions (Au3+ and Pd2+). We then investigated a few types of protein-rich biomass as adsorbents for precious metal ions. In the presence of various transition metal ions, Au3+ and Pd2+ were also selectively adsorbed onto the biomass tested. The bound precious metal ions were recovered by aqua regia after charring the metal-bound biomass. Finally, we demonstrated the successful recovery of Au3+ and Pd2+ from a metal refining solution and a metal plating waste using the biomass. We propose an environmentally friendly recycling system for precious metal ions using protein-rich biomass.. |
| 415. |
Tatsuo Maruyama, Hironari Matsushita, Yukiko Shimada, Ichiro Kamata, Misa Hanaki, Saori Sonokawa, Noriho Kamiya, Masahiro Goto, Proteins and protein-rich biomass as environmentally friendly adsorbents selective for precious metal ions, ENVIRONMENTAL SCIENCE & TECHNOLOGY, 10.1021/es061664x, 41, 4, 1359-1364, 2007.02, Proteins exhibit specific interactions with various metal ions, which play important roles in a living cell. Here, we found that various proteins selectively adsorbed precious metal ions at a wide range of pH values. Studies on protein sequences and on synthesized peptides revealed that a histidine-containing sequence had specific interactions with precious metal ions (Au3+ and Pd2+). We then investigated a few types of protein-rich biomass as adsorbents for precious metal ions. In the presence of various transition metal ions, Au3+ and Pd2+ were also selectively adsorbed onto the biomass tested. The bound precious metal ions were recovered by aqua regia after charring the metal-bound biomass. Finally, we demonstrated the successful recovery of Au3+ and Pd2+ from a metal refining solution and a metal plating waste using the biomass. We propose an environmentally friendly recycling system for precious metal ions using protein-rich biomass.. |
| 416. |
K. Nakashima, T. Maruyama, N. Kamiya, M. Goto, Activation of lipase in ionic liquids by modification with comb-shaped poly(ethylene glycol), Sci. Technol. Adv. Mater., vol.7, 692-698 (2006), 2007.01. |
| 417. |
J. Tominaga, Y. Kemori, Y. Tanaka, T. Maruyama, N. Kamiya, M. Goto, An enzymatic method for site-specific labeling of recombinant proteins with oligonucleotides, Chem. Commun., 401-403, 2007.01. |
| 418. |
Jo Tominaga, Yoshinori Kemori, Yusuke Tanaka, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, An enzymatic method for site-specific labeling of recombinant proteins with oligonucleotides, Chemical Communications, 10.1039/b613793h, 4, 401-403, 2007.01, DNA was site-specifically conjugated to a substrate peptide of microbial transglutaminase fused to the N- or C-terminus of target proteins without the loss of the proteins' functions of interest.. |
| 419. |
Yusuke Tanaka, Yukito Tsuruda, Motohiro Nishi, Noriho Kamiya, Masahiro Goto, Exploring enzymatic catalysis at a solid surface
A case study with transglutaminase-mediated protein immobilization, Organic and Biomolecular Chemistry, 10.1039/b701595j, 5, 11, 1764-1770, 2007, The factors affecting enzymatic protein immobilization with microbial transglutaminase (MTG) were explored. As model proteins, enhanced green fluorescent protein (EGFP) and glutathione S-transferase (GST) were chosen and tagged with a neutral Gln-donor substrate peptide for MTG (Leu-Leu-Gln-Gly, LLQG-tag) at their C-terminus. To create a specific surface, displaying reactive Lys residues, to be cross-linked with the Gln residue in the LLQG-tag of target proteins by MTG catalysis, a polystyrene surface was physically coated with β-casein. Both recombinant proteins were immobilized onto the β-casein-coated surface only in the presence of active MTG, indicating that those proteins were enzymatically immobilized to the surface. MTG-mediated protein immobilization markedly depends on the pH and ionic strength of the reaction media. The optimal pH range of MTG-mediated immobilization of both recombinant proteins was around 5, at which point the MTG-catalyzed reaction in aqueous solution is not normally preferred. By utilizing a pH-dependent change in EGFP fluorescence, we found that the apparent pH at the surface is likely to be lower than bulk pH, this difference is not attributed to an optimal pH shift in MTG-mediated immobilization. On the other hand, lower yields of protein immobilization at higher ionic strength suggest that electrostatic interaction is a key factor governing MTG catalysis at a solid surface. The results of this study indicate that, in enzymatic catalysis at a solid surface, the concentration of substrates at the surface can enhance the catalytic efficiency, and this could alter the pH dependence of enzymatic catalysis.. |
| 420. |
Hidehiko Hirakawa, Noriho Kamiya, Tsutomu Tanaka, Teruyuki Nagamune, Intramolecular electron transfer in a novel cytochrome P450cam system with a site-specific branched structure, 2006 AIChE Annual Meeting
2006 AIChE Annual Meeting, 2006.12. |
| 421. |
H. Piao, N. Kamiya, J. Watanabe, H. Yokoyama, A. Hirata, T. Fujii, I. Shimizu, S. Ito, M. Goto, Oral delivery of diclofenac sodium using a novel solid-in-oil suspension, Int. J. Pharm., vol.313, 159-162 (2006) , 2006.11. |
| 422. |
Kojiro Shimojo, Noriho Kamiya, Fumito Tani, Hirochika Naganawa, Yoshinori Naruta, Masahiro Goto, Extractive solubilization, structural change, and functional conversion of cytochrome c in ionic liquids via crown ether complexation, Analytical chemistry, 10.1021/ac0612877, 78, 22, 7735-7742, 2006.11, This article reports on the extraction behavior of heme proteins from an aqueous phase into ionic liquids (ILs) with dicyclohexano-18-crown-6 (DCH18C6), and the structure-function relationship of cytochrome c (Cyt-c) dissolved in ILs. We have found that DCH18C6 enables transfer of Lys-rich proteins into ILs via supramolecular complexation. The hydrophobicity and functional groups of ILs have a great influence on protein partitioning, and a hydroxyl group-containing IL with DCH18C6 is capable of the quantitative partitioning of Cyt-c. On the other hand, protein transfer using conventional organic solvents is negligibly small. UV-visible, CD, and resonance Raman spectroscopic characterizations indicate that the sixth ligand Met 80 in the heme group of the Cyt-c-DCH18C6 complex in IL is replaced by other amino acid residues of the peptide chain and that a non-natural, six-coordinate, low-spin ferric heme structure is induced in IL. Solubilization of Cyt-c in IL causes the environmental change of the heme vicinity of Cyt-c, which triggers the functional conversion of Cyt-c from an electron-transfer protein to peroxidase. The Cyt-c-DCH18C6 complex in IL provides remarkably high peroxidase activity compared with native Cyt-c, because of enhancement of the affinity for H2O2.. |
| 423. |
Kojiro Shimojo, Noriho Kamiya, Fumito Tani, Hirochika Naganawa, Yoshinori Naruta, Masahiro Goto, Extractive solubilization, structural change, and functional conversion of cytochrome c in ionic liquids via crown ether complexation, ANALYTICAL CHEMISTRY, 10.1021/ac0612877, 78, 22, 7735-7742, 2006.11, This article reports on the extraction behavior of heme proteins from an aqueous phase into ionic liquids (ILs) with dicyclohexano-18-crown-6 (DCH18C6), and the structure-function relationship of cytochrome c (Cyt-c) dissolved in ILs. We have found that DCH18C6 enables transfer of Lys-rich proteins into ILs via supramolecular complexation. The hydrophobicity and functional groups of ILs have a great influence on protein partitioning, and a hydroxyl group-containing IL with DCH18C6 is capable of the quantitative partitioning of Cyt-c. On the other hand, protein transfer using conventional organic solvents is negligibly small. UV-visible, CD, and resonance Raman spectroscopic characterizations indicate that the sixth ligand Met 80 in the heme group of the Cyt-c-DCH18C6 complex in IL is replaced by other amino acid residues of the peptide chain and that a non-natural, six-coordinate, low-spin ferric heme structure is induced in IL. Solubilization of Cyt-c in IL causes the environmental change of the heme vicinity of Cyt-c, which triggers the functional conversion of Cyt-c from an electron-transfer protein to peroxidase. The Cyt-c-DCH18C6 complex in IL provides remarkably high peroxidase activity compared with native Cyt-c, because of enhancement of the affinity for H2O2.. |
| 424. |
K. Nakashima, T. Maruyama, N. Kamiya, M. Goto, Homogeneous Enzymatic Reaction in Ionic Liquids with Poly(ethylene glycol)-Modified Subtilisin, Org. Biomol. Chem., vol.4, 3462-3467 (2006), 2006.10. |
| 425. |
K. Shimojo, N. Kamiya, F. Tani, H. Naganawa, Y. Naruta, M. Goto, Extractive Solubilization, Structural Change and Functional Conversion of Cytochrome c in Ionic Liquids via Crown Ether Complexation, Anal. Chem., vol.78, 7735-7742 (2006), 2006.10. |
| 426. |
K.Shimojo, K.Nakashima, N.Kamiya, M.Goto, Crown-ether mediated extraction and functional conversion of cytochrome c in ionic liquids, Biomacromolecules, vol.7, 2-5 (2006), 2006.10. |
| 427. |
Kazunori Nakashima, Jun Okada, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Activation of lipase in ionic liquids by modification with comb-shaped poly(ethylene glycol), Science and Technology of Advanced Materials, 10.1016/j.stam.2006.06.008, 7, 7, 692-698, 2006.10, Outstanding activation of an enzyme in ionic liquids (ILs) has been demonstrated by covalent modification with comb-shaped poly(ethylene glycol) (PEG) (PM13). Candida rugosa lipase modified with PM13 (PM13-CRL) was readily solubilized in all the ILs tested ([Emim][Tf2N], [C2OC1mim][Tf2N] and [C2OHmim][Tf2N]) containing 0.5% (v/v) of water, whereas native lipase did not dissolve in any of the ILs. The results for transesterification of 2-phenyl-1-propanol with vinyl acetate using lipase in ILs revealed that the PM13-CRL conjugate exhibits a high catalytic activity while suspended native lipase shows little activity. The hydrophobicity of ILs somewhat affected the enzyme activity and a more hydrophobic IL such as [Emim][Tf2N] was preferable for the lipase reaction, as was also observed in enzymatic reaction in conventional organic solvents. The enzyme activities in ILs were much higher than those in organic solvents, the excellent activity being associated with unique properties such as the hydrophobicity and the high polarity of ILs. Furthermore, the PM13--CRL conjugate exhibited a high storage stability in [Emim][Tf2N].. |
| 428. |
Kazunori Nakashima, Jun Okada, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Activation of lipase in ionic liquids by modification with comb-shaped poly(ethylene glycol), SCIENCE AND TECHNOLOGY OF ADVANCED MATERIALS, 10.1016/j.stam.2006.06.008, 7, 7, 692-698, 2006.10, Outstanding activation of an enzyme in ionic liquids (ILs) has been demonstrated by covalent modification with comb-shaped polytethylene glycol) (PEG) (PM13). Candida rugosa lipase modified with PM13 (PM13-CRL) was readily solubilized. in all the ILs tested ([Emim][Tf2N], [C(2)OC(1)mim][Tf2N] and [C(2)OHmim][Tf2N]) containing 0.5% (v/v) of water, whereas native lipase did not dissolve in any of the ILs. The results for transesterification of 2-phenyl-1-propanol with vinyl acetate using lipase in ILs revealed that the PM13-CRL conjugate exhibits a high catalytic activity while suspended native lipase shows little activity. The hydrophobicity of ILs somewhat affected the enzyme activity and a more hydrophobic IL such as [Emim][Tf2N] was preferable for the lipase reaction, as was also observed in enzymatic reaction in conventional organic solvents. The enzyme activities in ILs were much higher than those in organic solvents, the excellent activity being associated with unique properties such as the hydrophobicity and the high polarity of ILs. Furthermore, the PM13-CRL conjugate exhibited a high storage stability in [Emim][Tf2N], (c) 2006 NIMS and Elsevier Ltd. All rights reserved.. |
| 429. |
Tsuyoshi Mouri, Junji Michizoe, Hirofumi Ichinose, Noriho Kamiya, Masahiro Goto, A recombinant Escherichia coli whole cell biocatalyst harboring a cytochrome P450cam monooxygenase system coupled with enzymatic cofactor regeneration, Applied Microbiology and Biotechnology, 10.1007/s00253-005-0289-y, 72, 3, 514-520, 2006.09, A cytochrome P450cam monooxygenase (P450cam) system from the soil bacterium Pseudomonas putida requires electron transfer among three different proteins and a cofactor, nicotinamide adenine dinucleotide (NADH), for oxygenation of its natural substrate, camphor. Herein, we report a facile way to significantly enhance the catalytic efficiency of the P450cam system by the coupling of its native electron transfer system with enzymatic NADH regeneration catalyzed by glycerol dehydrogenase (GLD) in Escherichia coli whole cell biocatalysts. Recombinant E. coli harboring the P450cam system, but lacking GLD, exhibited little activity for camphor hydroxylation. In contrast, coexpression of GLD with the proteinaceous electron transfer components of P450cam resulted in about tenfold improvement in the substrate conversion, implying that the whole cell biocatalyst utilized molecular oxygen, endogenous NADH, and glycerol in the cell for catalysis. The addition of glycerol to the reaction media further promoted camphor hydroxylation, suggesting that exogenous glycerol is also available for GLD in the host cell and actively participates in the catalytic cycle. These results clearly show the utility of GLD towards functional reconstruction of the native P450cam system. The present approach may also be useful for E. coli whole cell biocatalysts with the other NADH-dependent oxygenases and oxidoreductases.. |
| 430. |
Kazunori Nakashima, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Homogeneous enzymatic reactions in ionic liquids with poly(ethylene glycol)-modified subtilisin, Organic and Biomolecular Chemistry, 10.1039/b608920h, 4, 18, 3462-3467, 2006.09, Subtilisin Carlsberg was covalently modified with comb-shaped poly(ethylene glycol) (PM13). PM13-modified subtilisin (PM 13-Sub) was readily solubilized in three different ionic liquids (ILs), i.e., [Emim][Tf2N], [C2OC1mim][Tf 2N] and [C2OHmim][Tf2N]. Analysis of homogeneous enzymatic reactions in the ILs revealed that PM13-Sub exhibited excellent catalytic performance while the native enzyme suspended in ILs showed no activity. Hydrophobicity of ILs slightly affected enzyme activity, and the relatively hydrophobic IL [Emim][Tf2N] was the preferred medium for enzymatic reactions, similar to enzymatic reactions in conventional organic solvents. Enzyme activity was much higher in [Emim][Tf2N] than in conventional organic solvents, and excellent activity was associated with unique properties of ILs such as hydrophobicity and high polarity. Furthermore, PM13-Sub showed good stability in [Emim][Tf 2N], and maintained 80% of its initial activity after 60 h.. |
| 431. |
Tsuyoshi Mouri, Noriho Kamiya, Masahiro Goto, Increasing the catalytic performance of a whole cell biocatalyst harboring a cytochrome P450cam system by stabilization of an electron transfer component, Biotechnology letters, 10.1007/s10529-006-9118-3, 28, 18, 1509-1513, 2006.09, Catalytic activity of a recombinant Escherichia coli whole cell biocatalyst harboring a cytochrome P450cam monooxygenase system from Pseudomonas putida coupled with enzymatic co-factor regeneration was investigated. About 0.7 μmol camphor was hydroxylated per mg dry cells at 4°C in 50 mM Tris/HCl buffer (pH 7.4) when utilizing a stable putidaredoxin (Pdx) mutant, C73S/C85S-Pdx (Cys73Ser, Cys85Ser double mutant), instead of wild-type Pdx, which was about two-fold improvement in the substrate conversion. Ten-micromole camphor was completely hydroxylated at 20°C in 6 h by 15 mg dry cell weight of whole cell biocatalyst including C73S/C85S-Pdx. Thus, modulation of protein-protein interaction in multicomponent enzymatic catalysis in whole cells is important.. |
| 432. |
Tsuyoshi Mouri, Junji Michizoe, Hirofumi Ichinose, Noriho Kamiya, Masahiro Goto, A recombinant Escherichia coli whole cell biocatalyst harboring a cytochrome P450cam monooxygenase system coupled with enzymatic cofactor regeneration, APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, 10.1007/s00253-005-0289-y, 72, 3, 514-520, 2006.09, A cytochrome P450cam monooxygenase (P450cam) system from the soil bacterium Pseudomonas putida requires electron transfer among three different proteins and a cofactor, nicotinamide adenine dinucleotide (NADH), for oxygenation of its natural substrate, camphor. Herein, we report a facile way to significantly enhance the catalytic efficiency of the P450cam system by the coupling of its native electron transfer system with enzymatic NADH regeneration catalyzed by glycerol dehydrogenase (GLD) in Escherichia coli whole cell biocatalysts. Recombinant E. coli harboring the P450cam system, but lacking GLD, exhibited little activity for camphor hydroxylation. In contrast, coexpression of GLD with the proteinaceous electron transfer components of P450cam resulted in about tenfold improvement in the substrate conversion, implying that the whole cell biocatalyst utilized molecular oxygen, endogenous NADH, and glycerol in the cell for catalysis. The addition of glycerol to the reaction media further promoted camphor hydroxylation, suggesting that exogenous glycerol is also available for GLD in the host cell and actively participates in the catalytic cycle. These results clearly show the utility of GLD towards functional reconstruction of the native P450cam system. The present approach may also be useful for E. coli whole cell biocatalysts with the other NADH-dependent oxygenases and oxidoreductases.. |
| 433. |
Tsuyoshi Mouri, Noriho Kamiya, Masahiro Goto, Increasing the catalytic performance of a whole cell biocatalyst harboring a cytochrome P450cam system by stabilization of an electron transfer component, BIOTECHNOLOGY LETTERS, 10.1007/s10529-006-9118-3, 28, 18, 1509-1513, 2006.09, Catalytic activity of a recombinant Escherichia coli whole cell biocatalyst harboring a cytochrome P450cam monooxygenase system from Pseudomonas putida coupled with enzymatic co-factor regeneration was investigated. About 0.7 mu mol camphor was hydroxylated per mg dry cells at 4 degrees C in 50 mM Tris/HCl buffer (pH 7.4) when utilizing a stable putidaredoxin (Pdx) mutant, C73S/C85S-Pdx (Cys73Ser, Cys85Ser double mutant), instead of wild-type Pdx, which was about two-fold improvement in the substrate conversion. Ten-micromole camphor was completely hydroxylated at 20 degrees C in 6 h by 15 mg dry cell weight of whole cell biocatalyst including C73S/C85S-Pdx. Thus, modulation of protein-protein interaction in multicomponent enzymatic catalysis in whole cells is important.. |
| 434. |
T. Maruyama, T. Shinohara, T. Hosogi, H. Ichinose, N. Kamiya, M. Goto, Masking oligonucleotides improve sensitivity of mutation detection based on guanine quenching, Anal. Biochem., vol.354, 8-14 (2006), 2006.07. |
| 435. |
Tatsuo Maruyama, Toshimitsu Shinohara, Takuya Hosogi, Hirofumi Ichinose, Noriho Kamiya, Masahiro Goto, Masking oligonucleotides improve sensitivity of mutation detection based on guanine quenching, Analytical Biochemistry, 10.1016/j.ab.2006.03.056, 354, 1, 8-14, 2006.07, Guanine quenching of a fluorescence-labeled DNA probe is a powerful tool for detecting a mutation in a targeted site of a DNA strand. However, a different guanine adjacent to a targeted site can interfere with detection of a point mutation, resulting in unsatisfactory sensitivity. In the current study, we developed a simple method to improve sensitivity of the guanine quenching method using a masking DNA oligonucleotide. The simple addition of a masking DNA oligonucleotide was found to mask the interference of a different guanine in a target oligonucleotide on fluorescence and to enhance difference in the quenching ratio between wild-type and mutant oligonucleotides. Based on this strategy, we succeeded in discriminating various mutations from the wild-type YMDD motif of the hepatitis B virus DNA polymerase gene using guanine quenching with a masking oligonucleotide.. |
| 436. |
T Maruyama, T Shinohara, T Hosogi, H Ichinose, N Kamiya, M Goto, Masking oligonucleotides improve sensitivity of mutation detection based on guanine quenching, ANALYTICAL BIOCHEMISTRY, 10.1016/j.ab.2006.03.056, 354, 1, 8-14, 2006.07, Guanine quenching of a fluorescence-labeled DNA probe is a powerful tool for detecting a mutation in a targeted site of a DNA strand. However, a different guanine adjacent to a targeted site can interfere with detection of a point mutation, resulting in unsatisfactory sensitivity. In the current study, we developed a simple method to improve sensitivity of the guanine quenching method using a masking DNA oligonucleotide. The simple addition of a masking DNA oligonucleotide was found to mask the interference of a different guanine in a target oligonucleotide on fluorescence and to enhance difference in the quenching ratio between wild-type and mutant oligonucleotides. Based on this strategy, we succeeded in discriminating various mutations from the wild-type YMDD motif of the hepatitis B virus DNA polymerase gene using guanine quenching with a masking oligonucteotide. (c) 2006 Elsevier Inc. All rights reserved.. |
| 437. |
T.Mouri, N.Kamiya, M.Goto, Increasing the catalytic performance of a whole cell biocatalyst harboring a cytochrome P450cam system by stabilization of an electron transfer component, Biotechnol. Lett., vol.28, 1509-1513 (2006), 2006.05. |
| 438. |
T.Mouri, J.Michizoe, H.Ichinose, N.Kamiya, M.Goto, A recombinant Escherichia coli whole cell biocatalyst harboring a cytochrome P450cam monooxygenase system coupled with enzymatic cofactor regeneration, Appl. Microbiol. Biotechnol., vol.72, 514-520 (2006), 2006.04. |
| 439. |
Hongyu Piao, Noriho Kamiya, Junji Watanabe, Hideakira Yokoyama, Akihiko Hirata, Takeru Fujii, Ichiro Shimizu, Susumu Ito, Masahiro Goto, Oral delivery of diclofenac sodium using a novel solid-in-oil suspension, International Journal of Pharmaceutics, 10.1016/j.ijpharm.2006.02.003, 313, 1-2, 159-162, 2006.04, The present work reports on a new pharmaceutical formulation for oral delivery of diclofenac sodium (DFNa), a non-steroidal anti-inflammatory drug (NSAID). Although DFNa itself is water-soluble at neutral pH, it was readily suspended in soybean oil via complex formation with an edible lipophilic surfactant and a matrix protein. The resulting solid-in-oil (S/O) suspension containing stably encapsulated DFNa in an oil phase markedly reduced the risks for gastrointestinal ulcers upon oral administration even at the LD50 level in rats (ca. 50 mg/kg DFNa). In addition, plasma concentration of DFNa upon administration of an S/O suspension was comparable with that of the aqueous counterpart at the same DFNa dose. These results indicate the potential use of S/O suspensions as novel oil-based pharmaceutical formulations for oral delivery of water-soluble drugs without causing severe mucitis.. |
| 440. |
H Piao, N Kamiya, J Watanabe, H Yokoyama, A Hirata, T Fujii, Shimizu, I, S Ito, M Goto, Oral delivery of diclofenac sodium using a novel solid-in-oil suspension, INTERNATIONAL JOURNAL OF PHARMACEUTICS, 10.1016/j.ijpharm.2006.02.003, 313, 1-2, 159-162, 2006.04, The present work reports on a new pharmaceutical formulation for oral delivery of diclofenac sodium (DFNa), a non-steroidal anti-inflammatory drug (NSAID). Although DFNa itself is water-soluble at neutral pH, it was readily suspended in soybean oil via complex formation with an edible lipophilic surfactant and a matrix protein. The resulting solid-in-oil (S/O) suspension containing stably encapsulated DFNa in an oil phase markedly reduced the risks for gastrointestinal ulcers upon oral administration even at the LD50 level in rats (ca. 50 mg/ka DFNa). In addition, plasma concentration of DFNa upon administration of an S/O suspension was comparable with that of the aqueous counterpart at the same DFNa dose. These results indicate the potential use of S/O suspensions as novel oil-based pharmaceutical formulations for oral delivery of water-soluble drugs without causing severe mucitis. (R) 2006 Elsevier B.V. All rights reserved.. |
| 441. |
Kojiro Shimojo, Kazunori Nakashima, Noriho Kamiya, Masahiro Goto, Crown ether-mediated extraction and functional conversion of cytochrome c in ionic liquids, Biomacromolecules, 10.1021/bm050847t, 7, 1, 2-5, 2006.01, We report that a macrocyclic ligand enables transfer of a protein from an aqueous phase to ionic liquids. The extraction behavior of heme protein cytochrome c (Cyt-c) from an aqueous phase into ionic liquids was investigated with crown ethers. A hydroxyl-group-containing ionic liquid with dicyclohexano-18-crown-6 was found to be capable of quantitative partitioning of Cyt-c, whereas the protein transfer using conventional organic solvents was negligibly small. Furthermore, we clarified that Cyt-c solubilized in ionic liquids caused a structural transformation of Cyt-c, which triggers its functional conversion from an electron-transfer protein to peroxidase.. |
| 442. |
Kojiro Shimojo, Kazunori Nakashima, Noriho Kamiya, Masahiro Goto, Crown ether-mediated extraction and functional conversion of cytochrome c in ionic liquids, Biomacromolecules, 10.1021/bm050847t, 7, 1, 2-5, 2006.01, We report that a macrocyclic ligand enables transfer of a protein from an aqueous phase to ionic liquids. The extraction behavior of heme protein cytochrome c (Cyt-c) from an aqueous phase into ionic liquids was investigated with crown ethers. A hydroxyl-group-containing ionic liquid with dicyclohexano-18-crown-6 was found to be capable of quantitative partitioning of Cyt-c, whereas the protein transfer using conventional organic solvents was negligibly small. Furthermore, we clarified that Cyt-c solubilized in ionic liquids caused a structural transformation of Cyt-c, which triggers its functional conversion from an electron-transfer protein to peroxidase. © 2006 American Chemical Society.. |
| 443. |
Solubilization and high activity of enzymes in ionic liquids by modification with comb-shaped poly (ethylene glycol)
Subtilisin Carlsberg was modified with comb-shaped poly (ethylene glycol), PM13. PM13 modified subtilisin (PM13-Sub) could be solubilized clearly in a wide range of pure ILs, whereas native subtilisin was not soluble in any ILs. Furthermore, PM13-Sub dissolved in ILs exhibited the excellent catalytic activity in transesterification of N-acetyl-L-phenylalanine ethyl ester with 1-butanol. This markedly high activity in the IL was superior to that in organic solvents commonly used for enzymatic catalysis. PM13-Sub was also found to retain its catalytic activity in the ILs for a prolonged period.. |
| 444. |
Hirofumi Ichinose, Momoko Kitaoka, Nobuko Okamura, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Detection of single-base mutations by fluorogenic ribonuclease protection assay, Analytical chemistry, 10.1021/ac050782k, 77, 21, 7047-7053, 2005.11, The ribonuclease protection assay is a generally applicable technique for the detection of known mutations. We have developed a simple and rapid method for mutation detection based on the ribonuclease protection assay using fluorescently labeled oligodeoxyribonucleotide probes. The fluorogenic ribonuclease protection (FRAP) assay uses two differently labeled oligodeoxyribonucleotides, a donor probe and an acceptor probe, to obtain a fluorescence resonance energy transfer (FRET) signal. We have utilized the FRAP assay for the detection of a single-base mutation in the YMDD motif of the hepatic B virus DNA polymerase gene. The occurrence of mismatch-selective RNA cleavage was successfully discriminated by measuring the FRET signal between the donor and acceptor probes. Moreover, mutation sensing was successfully visualized by a UV transillumination. This simple and rapid mutation sensing method should facilitate a high-throughput mutation analysis.. |
| 445. |
H Ichinose, M Kitaoka, N Okamura, T Maruyama, N Kamiya, M Goto, Detection of single-base mutations by fluorogenic ribonuclease protection assay, ANALYTICAL CHEMISTRY, 10.1021/ac050782k, 77, 21, 7047-7053, 2005.11, The ribonuclease protection assay is a generally applicable technique for the detection of known mutations. We have developed a simple and rapid method for mutation detection based on the ribonuclease protection assay using fluorescently labeled oligodeoxyribonucleotide probes. The fluorogenic ribonuclease protection (FRAP) assay uses two differently labeled oligodeoxyribonucleotides, a donor probe and an acceptor probe, to obtain a fluorescence resonance energy transfer (FRET) signal. We have utilized the FRAP assay for the detection of a single-base mutation in the YMDD motif of the hepatic B virus DNA polymerase gene. The occurrence of mismatch-selective RNA cleavage was successfully discriminated by measuring the FRET signal between the donor and acceptor probes. Moreover, mutation sensing was successfully visualized by a UV transillumination. This simple and rapid mutation sensing method should facilitate a high-throughput mutation analysis.. |
| 446. |
Eiichi Toorisaka, Masakazu Hashida, Noriho Kamiya, Hiroshige Ono, Yuko Kokazu, Masahiro Goto, An enteric-coated dry emulsion formulation for oral insulin delivery, Journal of Controlled Release, 10.1016/j.jconrel.2005.05.022, 107, 1, 91-96, 2005.09, A novel oral dosage formulation of insulin consisting of a surfactant, a vegetable oil, and a pH-responsive polymer has been developed. First, a solid-in-oil (S/O) suspension containing a surfactant-insulin complex was prepared. Solid-in-oil-in-water (S/O/W) emulsions were obtained by homogenizing the S/O suspension and the aqueous solution of hydroxypropylmethylcellulose phthalate (HPMCP). A microparticulate solid emulsion formulation was successfully prepared from the S/O/W emulsions by extruding them to an acidic aqueous solution, followed by lyophilization. The insulin release from the resultant dry emulsion responded to the change in external environment simulated by gastrointestinal conditions, suggesting that the new enteric-coated dry emulsion formulation is potentially applicable for the oral delivery of peptide and protein drugs.. |
| 447. |
Kazunori Nakashima, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Comb-shaped poly(ethylene glycol)-modified subtilisin Carlsberg is soluble and highly active in ionic liquids, Chemical Communications, 10.1039/b505479f, 34, 4297-4299, 2005.09, Subtilisin Carlsberg conjugated with comb-shaped poly-(ethylene glycol) was solubilized in common ionic liquids without adding water, and exhibited higher transesterification activity in an ionic liquid, [Emim][Tf2N], than in organic solvents commonly used for enzymatic biotransformation.. |
| 448. |
Tatsuo Maruyama, Toshimitsu Shinohara, Hirofumi Ichinose, Momoko Kitaoka, Nobuko Okamura, Noriho Kamiya, Masahiro Goto, Mutation detection in DNA oligonucleotides based on a guanine quenching method coupled with enzymatic digestion of single-stranded DNA, Biotechnology letters, 10.1007/s10529-005-3681-x, 27, 18, 1349-1354, 2005.09, Fluorescence quenching by guanine allows DNA hybridization to be monitored and any point mutations in oligonucleotides to be detected. However, fluorescence quenching is often affected by untargeted guanine located in a protruding end (single-strand DNA) of the probe-target DNA duplex resulting in an unsatisfactory sensitivity. In the present study, we used enzymatic digestion of the protruding end of a probe-target DNA duplex to avoid interference by untargeted guanine on fluorescence quenching for detection of a nucleobase mutation. Enzymatic digestion of the protruding end of the DNA duplex fully prevented interference by untargeted guanine, and produced a marked difference in the quenching ratios (36% for wild-type, and 0% for mutant).. |
| 449. |
E Toorisaka, M Hashida, N Kamiya, H Ono, Y Kokazu, M Goto, An enteric-coated dry emulsion formulation for oral insulin delivery, JOURNAL OF CONTROLLED RELEASE, 10.1016/j.jconrel.2005.05.022, 107, 1, 91-96, 2005.09, A novel oral dosage formulation of insulin consisting of a surfactant, a vegetable oil, and a pH-responsive polymer has been developed. First, a solid-in-oil (S/O) suspension containing a surfactant-insulin complex was prepared. Solid-in-oil-in-water (S/O/W) emulsions were obtained by homogenizing the S/O suspension and the aqueous solution of hydroxypropylmethylcellulose phthalate (HPMCP). A microparticulate solid emulsion formulation was successfully prepared from the S/O/W emulsions by extruding them to an acidic aqueous solution, followed by lyophilization. The insulin release from the resultant dry emulsion responded to the change in external environment simulated by gastrointestinal conditions, suggesting that the new enteric-coated dry emulsion formulation is potentially applicable for the oral delivery of peptide and protein drugs. (c) 2005 Elsevier B.V. All rights reserved.. |
| 450. |
T Maruyama, T Shinohara, H Ichinose, M Kitaoka, N Okamura, N Kamiya, M Goto, Mutation detection in DNA oligonucleotides based on a guanine quenching method coupled with enzymatic digestion of single-stranded DNA, BIOTECHNOLOGY LETTERS, 10.1007/s10529-005-3681-x, 27, 18, 1349-1354, 2005.09, Fluorescence quenching by guanine allows DNA hybridization to be monitored and any point mutations in oligonucleotides to be detected. However, fluorescence quenching is often affected by untargeted guanine located in a protruding end (single-strand DNA) of the probe-target DNA duplex resulting in an unsatisfactory sensitivity. In the present study, we used enzymatic digestion of the protruding end of a probe-target DNA duplex to avoid interference by untargeted guanine on fluorescence quenching for detection of a nucleobase mutation. Enzymatic digestion of the protruding end of the DNA duplex fully prevented interference by untargeted guanine, and produced a marked difference in the quenching ratios (36% for wild-type, and 0% for mutant).. |
| 451. |
Hirofumi Ichinose, Noriho Kamiya, Masahiro Goto, Enzymatic redox cofactor regeneration in organic media
Functionalization and application of glycerol dehydrogenase and soluble transhydrogenase in reverse micelles, Biotechnology Progress, 10.1021/bp0500765, 21, 4, 1192-1197, 2005.07, An enzymatic system for the regeneration of redox cofactors NADH and NADPH was investigated in nanostructural reverse micelles using bacterial glycerol dehydrogenase (GLD) and soluble transhydrogenase (STH). Catalytic conversion of NAD+ to NADH was realized in the sodium dioctylsulfosuccinate (AOT)/isooctane reverse micellar system harboring GLD and a sacrificial substrate, glycerol. The initial rate of NADH regeneration was enhanced by exogenous addition of ammonium sulfate into the reverse micelles, suggesting that NH4+ acts as a monovalent cationic activator. STH was successfully entrapped in the AOT/isooctane reverse micelles as well as GLD and was revealed to be capable of catalyzing the stoichiometric hydrogen transfer reaction between NADP+ and NADPH in reverse micelles. These results indicate that GLD and STH have potential for use in redox cofactor recycling in reverse micelles, which allows the use of catalytic quantities of NAD(P)H in organic media.. |
| 452. |
J Tominaga, N Kamiya, S Doi, H Ichinose, T Maruyama, M Goto, Design of a specific peptide tag that affords covalent and site-specific enzyme immobilization catalyzed by microbial transglutaminase, BIOMACROMOLECULES, 10.1021/bm050193o, 6, 4, 2299-2304, 2005.07, Transglutaminase-mediated site-specific and covalent immobilization of an enzyme to chemically modified a,garose was explored. Using Escherichia coli alkaline phosphatase (AP) as a model, two designed specific peptide tags containing a reactive lysine (Lys) residue with different length Gly-Ser linkers for microbial transglutaminase (MTG) were genetically attached to N- or C-termini. For solid support, agarose gel beads were chemically modified with beta-casein to display reactive glutamine (Gln) residues on the support surface. Recombinant APs were enzymatically and covalently immobilized to casein-grafted agarose beads. Immobilization by MTG markedly depended on either the position or the length of the peptide tags incorporated to AP, suggesting steric constraint upon enzymatic immobilization. Enzymatically immobilized AP showed comparable catalytic turnover (k(cat)) to the soluble counterpart and comparable operational stability with chemically immobilized AP. These results indicate that attachment of a suitable specific peptide tag to the right position of a target protein is crucial for MTG-inediated formulation of highly active immobilized proteins.. |
| 453. |
H Ichinose, N Kamiya, M Goto, Enzymatic redox cofactor regeneration in organic media: Functionalization and application of glycerol dehydrogenase and soluble transhydrogenase in reverse micelles, BIOTECHNOLOGY PROGRESS, 10.1021/bp0500765, 21, 4, 1192-1197, 2005.07, An enzymatic system for the regeneration of redox cofactors NADH and NADPH was investigated in nanostructural reverse micelles using bacterial glycerol dehydrogenase (GLD) and soluble transhydrogenase (STH). Catalytic conversion of NAD(+) to NADH was realized in the sodium dioetylsulfosuccinate (AOT)/isooctane reverse micellar system harboring GLD and a sacrificial substrate, glycerol. The initial rate of NADH regeneration was enhanced by exogenous addition of ammonium sulfate into the reverse micelles, suggesting that NH4+ acts as a monovalent cationic activator. STH was successfully entrapped in the AOT/isooctane reverse micelles as well as GLD and was revealed to be capable of catalyzing the stoichiometric hydrogen transfer reaction between NADP(+) and NADPH in reverse micelles. These results indicate that GLD and STH have potential for use in redox cofactor recycling in reverse micelles, which allows the use of catalytic quantities of NAD(P)H in organic media.. |
| 454. |
Y Michizoe, H Ichinose, N Kamiya, T Maruyama, M Goto, Biodegradation of phenolic environmental pollutants by a surfactant-laccase complex in organic media, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 10.1263/jbb.99.642, 99, 6, 642-647, 2005.06, Oxidative degradation of phenolic environmental pollutants in organic media was investigated using a laccase complexed with surfactants. The catalytic activity of the surfactant-laccase complex in isooctane was markedly enhanced by appropriately adjusting the water content of the reaction medium using reverse micelles. The surfactant-laccase complex showed little activity towards the oxidative reaction of bisphenol A in water-saturated isooctane (i.e., 0.0055% [v/v] water) while effectively catalyzed the same reaction in isooctane containing 4% (v/v) water, which is over the maximum water solubility. The latter system was homogeneous and was only achieved by the aid of reverse micelles. With respect to the oxidation of bisphenol A, two products, 4-isopropylphenol and 4-isopropenylphenol, were identified by gas chromatography-mass spectrometry (GC-MS) analyses, indicating the oxidative degradation of the bis-phenolic structure of bisphenol A. We also found that the surfactant-laccase complex turned out to handle other environmental pollutants, chlorophenols, by the simultaneous addition of water and a redox mediator into the reaction medium using reverse micelles.. |
| 455. |
Eijiro Miyako, Tatsuo Maruyama, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Optical resolution of various amino acids using a supported liquid membrane encapsulating a surfactant-protease complex, Langmuir, 10.1021/la046789z, 21, 10, 4674-4679, 2005.05, We have encapsulated a surfactant-protease complex (the main protease used being a-chymotrypsin) in an organic phase of a supported liquid membrane (SLM) for the optical resolution of various amino acids. L-Isomers of amino acids were enantioselectively permeated through the SLM. The mechanism of the amino acid permeation through the SLM was considered to be as follows; an L-amino acid was enantioselectively esterified with ethanol by a surfactant-protease complex encapsulated in the SLM, and the resulting L-amino acid ethyl ester dissolved into the organic phase of the SLM and diffused across the SLM. Another surfactant-α-chymotrypsin complex in the receiving phase catalyzed ester hydrolysis to produce the initial L-amino acid and ethanol, which are water-soluble. Thus, the L-amino acid was selectively transported to the receiving phase through the SLM on the basis of the molecular recognition of the surfactant-protease complex in the SLM. It was found that the catalytic activity and enantioselectivity of the surfactant-protease complex governed the permeate flux of amino acids and the enantiomeric excess in the membrane separation.. |
| 456. |
E Miyako, T Maruyama, F Kubota, N Kamiya, M Goto, Optical resolution of various amino acids using a supported liquid membrane encapsulating a surfactant-protease complex, LANGMUIR, 10.1021/la046789z, 21, 10, 4674-4679, 2005.05, We have encapsulated a surfactant-protease complex (the main protease used being α-chymotrypsin) in an organic phase of a supported liquid membrane (SLM) for the optical resolution of various amino acids. L-Isomers of amino acids were enantioselectively permeated through the SLM. The mechanism of the amino acid permeation through the SLM was considered to be as follows; an L-amino acid was enantioselectively esterified with ethanol by a surfactant-protease complex encapsulated in the SLM, and the resulting L-amino acid ethyl ester dissolved into the organic phase of the SLM and diffused across the SLM. Another surfactant-α-chymotrypsin complex in the receiving phase catalyzed ester hydrolysis to produce the initial L-amino acid and ethanol, which are water-soluble. Thus, the L-amino acid was selectively transported to the receiving phase through the SLM on the basis of the molecular recognition of the surfactant-protease complex in the SLM. It was found that the catalytic activity and enantioselectivity of the surfactant-protease complex governed the permeate flux of amino acids and the enantiomeric excess in the membrane separation.. |
| 457. |
Hidehiko Hirakawa, Noriho Kamiya, Yutaka Kawarabayashi, Teruyuki Nagamune, Log P effect of organic solvents on a thermophilic alcohol dehydrogenase, Biochimica et Biophysica Acta - Proteins and Proteomics, 10.1016/j.bbapap.2004.12.007, 1748, 1, 94-99, 2005.04, An alcohol dehydrogenase from the hyperthermophilic archaeon Aeropyrum pernix was activated by water-miscible organic solvents. This activation was influenced by the kind and the concentration of the added organic solvents. The kcat was increased by a factor of over ten when the mole fraction of acetonitrile was 0.1. This effect was large when organic solvents with large log P values were added. In fact, the kcat showed a strong positive correlation with the log P value of the mixed solvent at a constant mole fraction of water, while it was not affected by the kind of organic solvents added. Both the activation enthalpy and the entropy decreased with an increase in log P. The contribution of the activation enthalpy to the free energy of activation was larger than that of the activation entropy, and the free energy of activation decreased with an increase in log P.. |
| 458. |
Kosuke Araki, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Metal ion-selective membrane prepared by surface molecular imprinting, Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences, 10.1016/j.jchromb.2004.12.030, 818, 2, 141-145, 2005.04, Surface molecular imprinting was applied to the preparation of an ion-selective polymeric membrane for the first time. The use of acrylonitrile-butadiene rubber and a porous solid support in the polymer matrix resulted in improved flexibility and mechanical strength of the imprinted membrane. The asymmetric porous structure of the membrane was observed by scanning electron microscopy. The selectivity of the zinc(II)-imprinted membrane was evaluated by competitive adsorption and permeation studies. The imprinted membrane showed higher adsorption affinity and permeation selectivity towards the imprinted zinc ion than the non-imprinted counterpart. On the basis of the results obtained, the permeation mechanism of the metal ions was considered to be hopping of metal ions on the binding sites in the membranes.. |
| 459. |
Hirakawa, Hidehiko, Kamiya, Noriho, Kawarabayashi, Yutaka, Nagamune, Teruyuki, Activation of a Thermophilic Alcohol Dehydrogenase by Organic Solvents, Asian Pacific Confederation of Chemical Engineering congress program and abstracts, 10.11491/apcche.2004.0.435.0, 2005.04. |
| 460. |
H Hirakawa, N Kamiya, Y Kawarabayashi, T Nagamune, Log P effect of organic solvents on a thermophilic alcohol dehydrogenase, BIOCHIMICA ET BIOPHYSICA ACTA-PROTEINS AND PROTEOMICS, 10.1016/j.bbapap.2004.12.007, 1748, 1, 94-99, 2005.04, An alcohol dehydrogenase from the hyperthermophilic archaeon Aeropyrum pernix was activated by water-miscible organic solvents. This activation was influenced by the kind and the concentration of the added organic solvents. The k(cat) was increased by a factor of over ten when the mole fraction of acetonitrile was 0.1. This effect was large when organic solvents with large log P values were added. In fact, the k(cat) showed a strong positive correlation with the log P value of the mixed solvent at a constant mole fraction of water, while it was not affected by the kind of organic solvents added. Both the activation enthalpy and the entropy decreased with an increase in log P. The contribution of the activation enthalpy to the free energy of activation was larger than that of the activation entropy, and the free energy of activation decreased with an increase in log P. (c) 2004 Published by Elsevier B.V.. |
| 461. |
K Araki, T Maruyama, N Kamiya, M Goto, Metal ion-selective membrane prepared by surface molecular imprinting, JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES, 10.1016/j.jchromb.2004.12.030, 818, 2, 141-145, 2005.04, Surface molecular imprinting was applied to the preparation of an ion-selective polymeric membrane for the first time. The use of acrylonitrile-butadiene rubber and a porous solid support in the polymer matrix resulted in improved flexibility and mechanical strength of the imprinted membrane. The asymmetric porous structure of the membrane was observed by scanning electron microscopy. The selectivity of the zinc(H)-imprinted membrane was evaluated by competitive adsorption and permeation studies. The imprinted membrane showed higher adsorption affinity and permeation selectivity towards the imprinted zinc ion than the non-imprinted counterpart. On the basis of the results obtained, the permeation mechanism of the metal ions was considered to be hopping of metal ions on the binding sites in the membranes. (c) 2005 Elsevier B.V. All rights reserved.. |
| 462. |
T Tanaka, N Kamiya, T Nagamune, N-terminal glycine-specific protein conjugation catalyzed by microbial transglutaminase, FEBS LETTERS, 10.1016/j.febslet.2005.02.064, 579, 10, 2092-2096, 2005.04, Here, we report the N-terminal glycine (Gly) residue of a target protein can be a candidate primary amine for site-specific protein conjugation catalyzed by microbial transglutaminase (MTG) from Streptomyces mobaraensis. Gly5-enhanced green fluorescent protein (EGFP) (EGFP with five additional Gly residues at its N-terminus) was cross-linked with Myc-dihydrofolate reductase (DHFR) (DHFR with the myc epitope sequence at its N-terminus) to yield DHFR-EGFP heterodimers. The reactivities of additional peptidyl linkers were investigated and the results obtained suggested that at least three additional Gly residues at the N-terminus were required to yield the EGFP-DHFR heterodimeric form. Site-directed mutagenesis analysis revealed marked preference of MTG for amino acids adjacent to the N-terminal Gly residue involved in the protein conjugation. In addition, peptide-protein conjugation was demonstrated by MTG-catalyzed N-terminal Gly-specific modification of a target protein with the myc epitope peptide. (c) 2005 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.. |
| 463. |
Tatsuo Maruyama, Tomoaki Takata, Hirofumi Ichinose, Noriho Kamiya, Hiroyuki Kuma, Naotaka Hamasaki, Hiroyuki Morita, Masahiro Goto, Detection of point mutations in the HBV polymerase gene using a fluorescence intercalator in reverse micelles, Biotechnology Progress, 10.1021/bp0496474, 21, 2, 575-579, 2005.03, We report a novel and simple method for mutation detection in DNA oligonucleotides using a double-stranded DNA specific dye (SYBR Green I) in nanostructured molecular assemblies, called reverse micelles. The intercalation of SYBR Green I into the duplex DNA exhibits fluorescent emission in a CTAB/isooctane reverse micellar system as well as in an aqueous solution. We found marked differences in the fluorescence intensity between perfectly matched and mismatched 52-mer synthetic oligonucleotides, which were designed to contain the YMDD motif of the hepatitis B virus (HBV) polymerase gene, in a reverse micellar solution. Using this method, we successfully detected a mutation in PCR-amplified oligonucleotides of the HBV polymerase gene in sera of four patients with chronic hepatitis B. This detection method does not require DNA immobilization, chemical modification of DNA, or any special apparatus; it only needs a normal fluorescence spectrophotometer, an inexpensive dye, and just 10 pmol of sample DNA.. |
| 464. |
T Maruyama, T Takata, H Ichinose, N Kamiya, H Kuma, N Hamasaki, H Morita, M Goto, Detection of point mutations in the HBV polymerase gene using a fluorescence intercalator in reverse micelles, BIOTECHNOLOGY PROGRESS, 10.1021/bp0496474, 21, 2, 575-579, 2005.03, We report a novel and simple method for mutation detection in DNA oligonucleotides using a double-stranded DNA specific dye (SYBR Green 1) in nanostructured molecular assemblies, called reverse micelles. The intercalation of SYBR Green I into the duplex DNA exhibits fluorescent emission in a CTAB/isooctane reverse micellar system as well as in an aqueous solution. We found marked differences in the fluorescence intensity between perfectly matched and mismatched 52-mer synthetic oligonucleotides, which were designed to contain the YMDD motif of the hepatitis B virus (HBV) polymerase gene, in a reverse micellar solution. Using this method, we successfully detected a mutation in PCR-amplified oligonucleotides of the HBV polyrnerase gene in sera of four patients with chronic hepatitis B. This detection method does not require DNA immobilization, chemical modification of DNA, or any special apparatus; it only needs a normal fluorescence spectrophotometer, an inexpensive dye, and just 10 pmol of sample DNA.. |
| 465. |
Eijiro Miyako, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, A supported liquid membrane encapsulating a surfactant-lipase complex for the selective separation of organic acids, Chemistry - A European Journal, 10.1002/chem.200400691, 11, 4, 1163-1170, 2005.02, We have developed a novel, lipase-facilitated, supported liquid membrane (SLM) for the selective separation of organic acids by encapsulating a surfactant-lipase complex in the liquid membrane phase. This system exhibited a high transport efficiency for 3-phenoxypropionic acid and enabled the selective separation of organic acids due to the different solubilities of the acids in the organic phase and the variable substrate specificity of the surfactant-lipase complex in the liquid membrane phase. We found that various parameters, such as the amount of surfactant-lipase complex in the SLM, the lipase concentration in the receiving phase, and the ethanol concentration in the feed phase, affected the transport behavior of organic acids. The optimum conditions were 5 gL-1 of the surfactant-CRL complex in the SLM (CRL = lipase from Candida rugosa), 8 gL-1 of PPL in the receiving phase (PPL = lipase from porcine pancreas), and an ethanol concentration of 50 vol %. Furthermore, we achieved high enantioselective transport of (S)-ibuprofen attributable to the enantioselectivity of the surfactant-CRL complex.. |
| 466. |
E Miyako, T Maruyama, N Kamiya, M Goto, A supported liquid membrane encapsulating a surfactant-lipase complex for the selective separation of organic acids, CHEMISTRY-A EUROPEAN JOURNAL, 10.1002/chem.200400691, 11, 4, 1163-1170, 2005.02, We have developed a novel, lipase-facilitated, supported liquid membrane (SLM) for the selective separation of organic acids by encapsulating a surfactant-lipase complex in the liquid membrane phase. This system exhibited a high transport efficiency for 3-phenoxypropionic acid and enabled the selective separation of organic acids due to the different solubilities of the acids in the organic phase and the variable substrate specificity of the surfactant-lipase complex in the liquid membrane phase. We found that various parameters, such as the amount of surfactant-lipase complex in the SLM, the lipase concentration in the receiving phase, and the ethanol concentration in the feed phase, affected the transport behavior of organic acids. ne optimum conditions were 5 gL(-1) of the surfactant-CRL complex in the SLM (CRL=Iipase from Candida rugosa), 8 gL(-1) of PPL in the receiving phase (PPL=Iipase from porcine pancreas), and an ethanol concentration of 50 vol%. Furthermore, we achieved high enantioselective transport of (S)ibuprofen attributable to the enantioselectivity of the surfactant-CRL complex.. |
| 467. |
APCChE2004 : The 10th Congress of the Asian Pacific Confederation of Chemical Engineering. |
| 468. |
T.Tanaka, N. Kamiya, T.Nagamune, N-terminal glycine-specific protein conjugation catalyzed by microbial transglutaminase, FEBS Letter, 10.1016/j.febslet.2005.02.064, 579, 10, 2092-2096, vol.579, 2092-2096 (2005), 2005.01. |
| 469. |
N.Kamiya, S.Doi, J.Tominaga, H.Ichinose, M.Goto, Transglutaminase-mediated protein immobilization to casein nanolayers created on a plastic surface, Biomacromolecules, 10.1021/bm0494895, 6, 1, 35-38, 6, 35-38 (2005), 2005.01. |
| 470. |
J. Tominaga, N. Kamiya, S. Doi, H. Ichinose, T. Maruyama, M. Goto, Design of a specific peptide tag that affords covalent and site-specific enzyme immobilization catalyzed by microbial transglutaminase, Biomacromolecules, 10.1021/bm050193o, 6, 4, 2299-2304, vol.6, 2299 -2304 (2005), 2005.01. |
| 471. |
Junji Michizoe, Hirofumi Ichinose, Noriho Kamiya, Tatsuo Maruyama, Masahiro Goto, Biodegradation of phenolic environmental pollutants by a surfactant-laccase complex in organic media, Journal of Bioscience and Bioengineering, 10.1263/jbb.99.642, 99, 6, 642-647, 2005.01, Oxidative degradation of phenolic environmental pollutants in organic media was investigated using a laccase complexed with surfactants. The catalytic activity of the surfactant-laccase complex in isooctane was markedly enhanced by appropriately adjusting the water content of the reaction medium using reverse micelles. The surfactant-laccase complex showed little activity towards the oxidative reaction of bisphenol A in water-saturated isooctane (i.e., 0.0055% [v/v] water) while effectively catalyzed the same reaction in isooctane containing 4% (v/v) water, which is over the maximum water solubility. The latter system was homogeneous and was only achieved by the aid of reverse micelles. With respect to the oxidation of bisphenol A, two products, 4-isopropylphenol and 4-isopropenylphenol, were identified by gas chromatography-mass spectrometry (GC-MS) analyses, indicating the oxidative degradation of the bis-phenolic structure of bis-phenol A. We also found that the surfactant-laccase complex turned out to handle other environmental pollutants, chlorophenols, by the simultaneous addition of water and a redox mediator into the reaction medium using reverse micelles.. |
| 472. |
Junji Michizoe, Hirofumi Ichinose, Noriho Kamiya, Tatsuo Maruyama, Masahiro Goto, Functionalization of the cytochrome P450cam monooxygenase system in the cell-like aqueous compartments of water-in-oil emulsions, Journal of Bioscience and Bioengineering, 10.1263/jbb.99.12, 99, 1, 12-17, 2005.01, The functionalization of the cytochrome P450cam monooxygenase system, which requires electron transfer among three different proteins, was investigated in the micro-scale aqueous compartments of stable water-in-oil (W/O) emulsions formed with the nonionic surfactant tetraethylene glycol dodecyl ether. Neither an organic-aqueous biphasic system nor a non-emulsified organic-aqueous solution containing the same amount of surfactant showed substantial hydroxylation of camphor, a natural substrate of P450cam, whereas substantial monooxygenation activity was detected when stable aqueous compartments were provided by the formation of W/O emulsions. Since the camphor hydroxylation in W/O emulsions was modest, we explored the integration of an enzymatic NADH regeneration system in order to effectively provide a reducing equivalent. Two different dehydrogenases, bacterial glycerol dehydrogenase (GLD) and yeast alcohol dehydrogenase (ADH), were selected, and each of these was coupled with the P450cam catalytic cycle in W/O emulsions. As a result, the camphor hydroxylation rate was successfully improved by approximately 5-fold when GLD was employed under optimized conditions. These results reveal the potential utility of the micro-scale cell-like aqueous compartments of W/O emulsions for multicomponent enzymatic reactions especially for substrates with low aqueous solubility.. |
| 473. |
J Michizoe, H Ichinose, N Kamiya, T Maruyama, M Goto, Functionalization of the cytochrome P450cam monooxygenase system in the cell-like aqueous compartments of water-in-oil emulsions, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 10.1263/jbb.99.012, 99, 1, 12-17, 2005.01, The functionalization of the cytochrome P450cam monooxygenase system, which requires electron transfer among three different proteins, was investigated in the micro-scale aqueous compartments of stable water-in-oil (W/O) emulsions formed with the nonionic surfactant tetraethylene glycol dodecyl ether. Neither an organic-aqueous biphasic system nor a non-emulsified organic-aqueous solution containing the same amount of surfactant showed substantial hydroxylation of camphor, a natural substrate of P450cam, whereas substantial monooxygenation activity was detected when stable aqueous compartments were provided by the formation of W/O emulsions. Since the camphor hydroxylation in W/O emulsions was modest, we explored the integration of an enzymatic NADH regeneration system in order to effectively provide a reducing equivalent. Two different dehydrogenases, bacterial glycerol dehydrogenase (GLD) and yeast alcohol dehydrogenase (ADH), were selected, and each of these was coupled with the P450cam catalytic cycle in W/O emulsions. As a result, the camphor hydroxylation rate was successfully improved by approximately 5-fold when GLD was employed under optimized conditions. These results reveal the potential utility of the micro-scale cell-like aqueous compartments of W/O emulsions for multicomponent enzymatic reactions especially for substrates with low aqueous solubility.. |
| 474. |
N Kamiya, S Doi, J Tominaga, H Ichinose, M Goto, Transglutaminase-mediated protein immobilization to casein nanolayers created on a plastic surface, BIOMACROMOLECULES, 10.1021/bm0494895, 6, 1, 35-38, 2005.01, An enzymatic method for covalent and site-specific immobilization of recombinant proteins on a plastic surface was explored. Using Escherichia coli alkaline phosphatase (AP) with a specific peptide tag (MKHKGS) genetically incorporated at the N-terminus as a model (NK-AP), microbial transglutaminase (MTG)-mediated protein immobilization was demonstrated. To generate a reactive surface for MTG, a 96-well polystyrene microtiter plate was physically coated with casein, a good MTG substrate. Successful immobilization of recombinant AP to the nanolayer of casein on the surface of the microtiter plate was verified by the detection of enzymatic activity. Since little activity was observed when wild-type AP was used, immobilization of NK-AP was likely directed by the specific peptide tag. When polymeric casein prepared by MTG was used as a matrix on the plate, the loading capacity of AP was increased about 2-fold compared to when casein was used as the matrix. Transglutaminase-mediated site-specific posttranslational modification of proteins offers one way of generating a variety of protein-based solid formulations for biotechnological applications.. |
| 475. |
Jo Tominaga, Noriho Kamiya, Satoshi Doi, Hirofumi Ichinose, Masahiro Goto, An enzymatic strategy for site-specific immobilization of functional proteins using microbial transglutaminase, Enzyme and Microbial Technology, 10.1016/j.enzmictec.2004.08.014, 35, 6-7, 613-618, 2004.12, A novel strategy for site-specific immobilization of recombinant proteins was investigated using microbial transglutaminase (MTG). Alkaline phosphatase (AP) was selected as a model protein and tagged with a short peptide (MKHKGS) at the N-terminus to provide a reactive Lys residue for MTG. On the other hand, casein, a well-known substrate for MTG, was chemically attached onto a polyacrylic resin to provide reactive Gln residues for the enzymatic immobilization of the recombinant AP. As a result, we succeeded in MTG-mediated functional immobilization of the recombinant AP onto casein-coated polyacrylic resin. It was found that the immobilized AP prepared using MTG exhibited much higher specific activity than that prepared by chemical modification. Moreover, enzymatic immobilization gave an immobilized formulation with higher stability upon repeated use than that obtained by physical adsorption. Use of this ability of MTG in posttranslational protein modification will provide us with a benign, site-specific immobilization method for functional proteins.. |
| 476. |
J Tominaga, N Kamiya, S Doi, H Ichinose, M Goto, An enzymatic strategy for site-specific immobilization of functional proteins using microbial transglutaminase, ENZYME AND MICROBIAL TECHNOLOGY, 10.1016/j.enzmictec.2004.08.014, 35, 6-7, 613-618, 2004.12, A novel strategy for site-specific immobilization of recombinant proteins was investigated using microbial transglutarninase (MTG). Alkaline phosphatase (A-P) was selected as a model protein and tagged with a short peptide (MKHKGS) at the N-terminus to provide a reactive Lys residue for MTG. On the other hand, casein, a well-known substrate for MTG, was chemically attached onto a polyacrylic resin to provide reactive Gln residues for the enzymatic immobilization of the recombinant AP. As a result, we succeeded in MTG-mediated functional immobilization of the recombinant AP onto casein-coated polyacrylic resin. It was found that the immobilized AP prepared using MTG exhibited much higher specific activity than that prepared by chemical modification. Moreover, enzymatic immobilization gave an immobilized formulation with higher stability upon repeated use than that obtained by physical adsorption. Use of this ability of MTG in posttranslational protein modification will provide us with a benign, site-specific immobilization method for functional proteins. (C) 2004 Elsevier Inc. All rights reserved.. |
| 477. |
Poly(ethylene glycol)-lipase complex highly active in ionic liquids
The alcoholysis catalyzed by the poly(ethylene glycol) (PEG)-lipase complex in ionic liquids was investigated. The alcoholysis between vinyl cinnamete and benzyl alcohol was catalyzed by lyophilized native lipase PS or the PEG-lipase PS complex in [Hmin][PF6]. The alcoholysis reaction involves acylation of the lipase and esterification of benzyl alcohol. The results show that the differences in activation among lipases was related to the solvent tolerance of the lipase or the dispersion of the PEG-lipase complexes.. |
| 478. |
Masafumi Sakono, Yu Mi Kawashima, Hirofumi Ichinose, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Direct refolding of inclusion bodies using reversed micelles, Biotechnology Progress, 10.1021/bp049887j, 20, 6, 1783-1787, 2004.11, The protein refolding of inclusion bodies was investigated using reversed micelles formed by aerosol OT (AOT). Ribonuclease A (RNase A) was overexpressed in Escherichia coli and used as native inclusion bodies. The enzymatic activity of RNase A was completely regained from the inclusion bodies within 14 h by solubilization in reversed micelles. To further enhance the refolding rate, a molecular chaperone, GroEL, was incorporated into the refolding system. The resultant refolding system including GroEL showed better performance under optimized conditions for the refolding of RNase A inclusion bodies. The refolding rate was considerably improved by the addition of the molecular chaperone, and the refolding step was completed in 1 h. The protein refolding in the GroEL-containing refolding system was strongly dependent on the coexistence of ATP and Mg2+, suggesting that the GroEL hosted in the reversed micelles was biologically active and assisted in the renaturation of the inclusion bodies. The addition of cold acetone to the reversed micellar solution allowed over 90% recovery of the renatured RNase A.. |
| 479. |
Tatsuo Maruyama, Tomoaki Kaji, Tomohiro Ohkawa, Ken Ichiro Sotowa, Hironari Matsushita, Fukiko Kubota, Noriho Kamiya, Katsuki Kusakabe, Masahiro Goto, Intermittent partition walls promote solvent extraction of metal ions in a microfluidic device, Analyst, 10.1039/b411630p, 129, 11, 1008-1013, 2004.11, Liquid-liquid extraction of metal ions was carried out in a microfluidic device which had intermittent partition walls in the center of the confluent microchannel 100 μm wide, 20 μm deep and 3 cm long. The intermittent partition walls (50 μm long) stabilized a two-phase (n-heptane-water) flow and allowed clear phase separation at the end-junction of the microchannel. Using this two-phase flow in the microchannel, yttrium ions were successfully extracted in a complex form with an extractant PC-88A (2-ethylhexyl phosphonic acid mono-2-ethylhexyl ester) from a feed aqueous phase to a n-heptane phase within a contact time of 1.5 s. Although the apparent interfacial area in the microchannel was reduced by introducing the partition walls, the presence of the partition walls improved the extraction efficiency 2-3 fold at a contact time of 0.12-0.24 s. Flow analyses using fluorescent beads and a computational fluidic dynamics simulation revealed that the partition walls induced a slight turbulence in the two-phase flow in the microchannel. This slight turbulence would result in the mixing of the aqueous phase and promote the transport of yttrium ions from the aqueous feed phase to the organic extractant phase.. |
| 480. |
Lian Chun Park, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Hiroyuki Kuma, Naotaka Hamasaki, Mutation detection in the drug-resistant hepatitis B virus polymerase gene using nanostructured reverse micelles, analytical sciences, 10.2116/analsci.20.1609, 20, 11, 1609-1611, 2004.11, The emergence of drug-resistant hepatitis B virus (HBV) has been reported in patients with prolonged administration of lamivudine, which is a potent drug for the prevention of HBV infection. Lamivudine-resistant HBV has several types of mutations at the YMDD motif of its DNA polymerase. We successfully demonstrated that monitoring the hybridization behavior in nanostructured reverse micelles enables us to detect single nucleotide polymorphisms (SNPs). With the aid of reverse micelles, a model 40-mer oligonucleotide containing a single-base substitution was clearly distinguished from the normal, complementary oligonucleotide. In addition, we extended this technique to a high-throughput analysis. The results obtained with a 96-well micro-plate reader indicated the possibility of SNPs detection toward multiple samples of patients.. |
| 481. |
M Sakono, YM Kawashima, H Ichinose, T Maruyama, N Kamiya, M Goto, Direct refolding of inclusion bodies using reversed micelles, BIOTECHNOLOGY PROGRESS, 10.1021/bp049887j, 20, 6, 1783-1787, 2004.11, The protein refolding of inclusion bodies was investigated using reversed micelles formed by aerosol OT (AOT). Ribonuclease A (RNase A) was overexpressed. in Escherichia coli and used as native inclusion bodies. The enzymatic activity of RNase A was completely regained from the inclusion bodies within 14 h by solubilization in reversed micelles. To further enhance the refolding rate, a molecular chaperone, GroEL, was incorporated into the refolding system. The resultant refolding system including GroEL showed better performance under optimized conditions for the refolding of RNase A inclusion bodies. The refolding rate was considerably improved by the addition of the molecular chaperone, and the refolding step was completed in 1 h. The protein refolding in the GroEL-containing refolding system was strongly dependent on the coexistence of ATP and Mg2+, suggesting that the GroEL hosted in the reversed micelles was biologically active and assisted in the renaturation of the inclusion bodies. The addition of cold acetone to the reversed micellar solution allowed over 90% recovery of the renatured RNase A.. |
| 482. |
LC Park, T Maruyama, N Kamiya, M Goto, H Kuma, N Hamasaki, Mutation detection in the drug-resistant hepatitis B virus polymerase gene using nanostructured reverse micelles, ANALYTICAL SCIENCES, 10.2116/analsci.20.1609, 20, 11, 1609-1611, 2004.11, The emergence of drug-resistant hepatitis B virus (HBV) has been reported in patients with prolonged administration of lamivudine, which is a potent drug for the prevention of HBV infection. Lamivudine-resistant HBV has several types of mutations at the YMDD motif of its DNA polymerase. We successfully demonstrated that monitoring the hybridization behavior in nanostructured reverse micelles enables us to detect single nucleotide polymorphisms (SNPs). With the aid of reverse micelles, a model 40-mer oligonucleotide containing a single-base substitution was clearly distinguished from the normal, complementary oligonucleotide. In addition, we extended this technique to a high-throughput analysis. The results obtained with a 96-well micro-plate reader indicated the possibility of SNPs detection toward multiple samples of patients.. |
| 483. |
有機酸の選択的分離が可能なリパーゼ-オルガノゲル膜システムの開発. |
| 484. |
逆ミセルによるインクルージョンボディの高効率リフォールディング. |
| 485. |
Tatsuo Maruyama, Hironari Matsushita, Jun Ichi Uchida, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Liquid membrane operations in a microfluidic device for separation of metal ions, Analytical Chemistry, 10.1021/ac049844h, 76, 15, 4495-4500, 2004.08, A three-phase flow, water/n-heptane/water, was constructed in a microchannel (100-μm width, 25-μm depth) on a glass microchip (3 cm × 7 cm) and was used as a liquid membrane for separation of metal ions. Surface modification of the microchannel by octadecylsilane groups induced spontaneous phase separation of the three-phase flow in the microfluidic device, which allows control of interfacial contact time and off-chip analysis using conventional analytical apparatus. Prior to the selective transport of a metal ion through the liquid membrane in the microchannel, the forward and backward extraction of yttrium and zinc ions was investigated in a two-phase flow on a microfluidic device using 2-ethylhexyl phosphonic acid mono-2-ethylhexyl ester (commercial name, PC-88A) as an extractant. The extraction conditions (contact time of the two phases, pH, extractant concentration) in the microfluidic device were examined. These investigations demonstrated that the conventional methodology for solvent extraction of metal ions is applicable to solvent extraction in a microchannel. Finally, we employed the three-phase flow in the microchannel as a liquid membrane and observed the selective transport of Y ion through the liquid membrane. In the present study, we succeeded, for the first time, in the selective separation of a targeted metal ion from an aqueous feed solution to a receiving phase within a few seconds by employing a liquid membrane formed in a microfluidic device.. |
| 486. |
T Maruyama, H Matsushita, J Uchida, F Kubota, N Kamiya, M Goto, Liquid membrane operations in a microfluidic device for selective separation of metal ions, ANALYTICAL CHEMISTRY, 10.1021/ac049844h, 76, 15, 4495-4500, 2004.08, A three-phase flow, water/n-heptane/water, was constructed in a microchannel (100-mum width, 25-mum depth) on a glass microchip (3 cm x 7 cm) and was used as a liquid membrane for separation of metal ions. Surface modification of the microchannel by octadecylsilane groups induced spontaneous phase separation of the three-phase flow in the microfluidic device, which allows control of interfacial contact time and off chip analysis using conventional analytical apparatus. Prior to the selective transport of a metal ion through the liquid membrane in the microchannel, the forward and backward extraction of yttrium and zinc ions was investigated in a two-phase flow on a microfluidic device using 2-ethylhexyl phosphonic acid mono-2-ethylhexyl ester (commercial name, PC-88A) as an extractant. The extraction conditions (contact time of the two phases, pH, extractant concentration) in the microfluidic device were examined. These investigations demonstrated that the conventional methodology for solvent extraction of metal ions is applicable to solvent extraction in a microchannel. Finally, we employed the three-phase flow in the microchannel as a liquid membrane and observed the selective transport of Y ion through the liquid membrane. In the present study, we succeeded, for the first time, in the selective separation of a targeted metal ion from an aqueous feed solution to a receiving phase within a few seconds by employing a liquid membrane formed in a microfluidic device.. |
| 487. |
Masafumi Sakono, Yu Mi Kawashima, Hirofumi Ichinose, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Efficient refolding of inclusion bodies by reversed micelles, kagaku kogaku ronbunshu, 10.1252/kakoronbunshu.30.468, 30, 4, 468-473, 2004.07, We investigated the refolding of protein in inclusion bodies using reversed micelles formed by aerosol OT (AOT). RNase A was overexpressed in E. coli and used to prepare inclusion bodies of RNase A, which were then subjected to protein refolding by the AOT/isooctane reversed micellar system. The RNase A was renatured in reversed micelles, and the refolding yield reached 100% in 18h. The addition of cold acetone to the reversed micellar solution allowed the recovery of the renatured RNase A without any loss of its activity. In the reversed micellar refolding system, the refolding conditions for inclusion bodies agreed with those for wild-type RNase A artificially denatured with a denaturant. This result indicates that the refolding mechanism of the inclusion bodies is similar to that of the denatured wild-type RNase A. A dilution method for refolding of the inclusion bodies caused the re-aggregation of RNase A, and only 40% of the native activity was regained. Reversed micelles effectively promoted the refolding of RNase A from its inclusion body, suggesting that a reversed micellar protein refolding method can yield a higher rate of renaturation than the conventional dilution method.. |
| 488. |
Eijiro Miyako, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Highly enantioselective separation using a supported liquid membrane encapsulating surfactant-enzyme complex, Journal of the American Chemical Society, 10.1021/ja049378d, 126, 28, 8622-8623, 2004.07, We developed a highly enantioselective separation system for the optically active compounds, (S)-ibuprofen and L-phenylalanine, from their racemic mixtures by employing a supported liquid membrane (SLM) encapsulating a surfactant-lipase complex (or a surfactant-α-chymotrypsin complex). In the present system, enzymes encapsulated in the liquid-membrane phase effectively drove the enantioselective transport of optically active compounds through the SLM. This novel SLM allowed high enantioselectivity (ee over 91%) in the optical resolution of racemic ibuprofen and phenylalanine.. |
| 489. |
Masafumi Sakono, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Refolding of denatured carbonic anhydrase B by reversed micelles formulated with nonionic surfactant, Biochemical Engineering Journal, 10.1016/j.bej.2004.01.003, 19, 3, 217-220, 2004.07, We have succeeded in the efficient refolding of denatured carbonic anhydrase B (CAB) using reversed micelles formulated with nonionic surfactant. The reversed micelles with nonionic tetraethylene glycol dodecyl ether prevented denatured CAB from aggregation in the refolding process, allowing a refolding yield of over 70% in 20 h, while the refolding yield of CAB with AOT/isooctane reversed micelle was only about 5%. These results demonstrated that the reversed micelles-mediated protein refolding technique could be extended to the protein which has strong interaction with AOT by simply altering the type of surfactant.. |
| 490. |
J Michizoe, Y Uchimura, H Ichinose, T Maruyama, N Kamiya, H Wariishi, S Furusaki, M Goto, Activation of manganese peroxidase in an organic medium using a mediator, BIOCHEMICAL ENGINEERING JOURNAL, 10.1016/j.bej.2003.09.008, 19, 1, 43-46, 2004.07, We found that both unsaturated fatty acids (UFAs) and 1-hydroxybenzotriazole (HBT) enhance the enzymatic activity of manganese peroxidase in an organic medium. The effects of hydrophobic unsaturated fatty acids directly dissolved in an organic medium and hydrophilic HBT encapsulated in reverse micelles on the oxidation activity of a surfactant-manganese peroxidase (MnP) complex were investigated. The addition of UFAs or HBT (mediator) using reverse micelles improved the oxidation of 2,4-dichlorophenol in toluene up to 3-fold the oxidation without each mediator. This study presents, for the first time, the possibility of MnP-catalyzed oxidation coupled with mediators in organic media. The capability of the latter system using HBT was further assessed by the oxidative conversion of environmental pollutants, i.e. 2.4-dichlorophenol, 2,4,5-trichlorophenol, 2,4,6-trichlorophenol, and bisphenol A (100, 84, 91, and 100% conversions at 12 h of reaction, respectively). (C) 2003 Elsevier B.V. All rights reserved.. |
| 491. |
E Miyako, T Maruyama, N Kamiya, M Goto, Highly enantioselective separation using a supported liquid membrane encapsulating surfactant-enzyme complex, JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, 10.1021/ja049378d, 126, 28, 8622-8623, 2004.07, We developed a highly enantioselective separation system for the optically active compounds, (S)-ibuprofen and L-phenylalanine, from their racemic mixtures by employing a supported liquid membrane (SLM) encapsulating a surfactant-lipase complex (or a surfactant-α-chymotrypsin complex). In the present system, enzymes encapsulated in the liquid-membrane phase effectively drove the enantioselective transport of optically active compounds through the SLM. This novel SLM allowed high enantioselectivity (ee over 91%) in the optical resolution of racemic ibuprofen and phenylalanine. Copyright © 2004 American Chemical Society.. |
| 492. |
M Sakono, T Maruyama, N Kamiya, M Goto, Refolding of denatured carbonic anhydrase B by reversed micelles formulated with nonionic surfactant, BIOCHEMICAL ENGINEERING JOURNAL, 10.1016/j.bej.2004.01.003, 19, 3, 217-220, 2004.07, We have succeeded in the efficient refolding of denatured carbonic anhydrase B (CAB) using reversed micelles formulated with nonionic surfactant. The reversed micelles with nonionic tetraethylene glycol dodecyl ether prevented denatured CAB from aggregation in the refolding process, allowing a refolding yield of over 70% in 20 h, while the refolding yield of CAB with AOT/isooctane reversed micelle was only about 5%. These results demonstrated that the reversed micelles-mediated protein refolding technique could be extended to the protein which has strong interaction with AOT by simply altering the type of surfactant. (C) 2004 Elsevier B.V. All rights reserved.. |
| 493. |
H Ichinose, J Michizoe, T Maruyama, N Kamiya, M Goto, Electron-transfer reactions and functionalization of cytochrome P450cam monooxygenase system in reverse micelles, LANGMUIR, 10.1021/la049752n, 20, 13, 5564-5568, 2004.06, Enzyme-based electron-transfer reactions involved in the cytochrome P450 monooxygenase system were investigated in nanostructural reverse micelles. A bacterial flavoprotein, putidaredoxin reductase (PdR), was activated and shown to be capable of catalyzing the electron transport from NADH to electron-carrier proteins such as cytochrome b(5) (tCyt-b5) and putidaredoxin (Pdx) in reverse micelles. Ferric tCyt-b5 in reverse micelles was effectively converted to its ferrous form by the exogenous addition of separately prepared reverse micellar solution harboring PdR and NADH. The fact that direct interactions of macromolecular proteins should be possible in the reverse micellar system encouraged us to functionalize a multicomponent monooxygenase system composed of the bacterial cytochrome P450cam (P450cam), putidaredoxin (Pdx), and PdR in reverse micelles. The successful camphor hydroxylation reaction catalyzed by P450cam was significantly dependent on the coexistence of Pdx, PdR, and NADH but not H2O2, suggesting that the oxygen-transfer reactions proceeded via a "monooxygenation" mechanism. This is the first report of a multicomponent cytochrome P450 system exhibiting enzymatic activity in organic media.. |
| 494. |
Takeshi Takazawa, Noriho Kamiya, Hiroshi Ueda, Teruyuki Nagamune, Enzymatic labeling of a single chain variable fragment of an antibody with alkaline phosphatase by microbial transglutaminase, Biotechnology and Bioengineering, 10.1002/bit.20019, 86, 4, 399-404, 2004.05, Functional cross-linking of a single chain Fv fragment of anti-hen egg-white lysozyme antibody (scFv) and alkaline phosphatase (AP) was explored using microbial transglutaminase (MTG) from Streptomyces mobaraensis. A specific peptidyl linker for MTG was genetically fused to the N-terminus of each protein and the resultant proteins were obtained separately by bacterial expression. The recombinant peptide-tagged scFv and AP were site-specifically cross-linked by MTG through the extra peptidyl linkers in vitro, which mainly yielded the heterodimer (i.e., scFv-AP conjugate). The enzymatic cross-linking reaction had little influence on either the antigen-binding ability of the scFv moiety or the enzymatic activity of the AP moiety of the conjugate, allowing use within an enzyme-linked immunosorbent assay. The results obtained suggest that the enzymatic approach with MTG facilitates the posttranslational construction of functional fusion proteins.. |
| 495. |
Tsutomu Tanaka, Noriho Kamiya, Teruyuki Nagamune, Peptidyl linkers for protein heterodimerization catalyzed by microbial transglutaminase, Bioconjugate Chemistry, 10.1021/bc034209o, 15, 3, 491-497, 2004.05, Specific peptidyl linkers that result in the heterodimerization of functional proteins, which is catalyzed by microbial transglutaminase from Streptomyces mobaraensis (MTG), were generated based on a ribonuclease S-peptide using site-directed mutagenesis. The peptidyl linkers designated as Lys-tag and Gln-tag were designed to possess sole reactive Lys or Gln residue that was amenable for selective Lys-Gln cross-linkage of different proteins. Green fluorescent protein variants, ECFP and EYFP, were employed as model proteins, and those Lys- and Gln-tags were fused to the N-termini of ECFP and EYFP, respectively. As a result, we succeeded in solely obtaining the ECFP-EYFP heterodimer without forming multiply cross-linked byproducts. It was found that the reactivity of peptidyl linkers varied according to the type of amino acid to be replaced. Peptidyl linkers with a basic amino acid (Arg) exhibited the highest reactivity in the cross-linking reaction, suggesting the cationic residue substrate preference of MTG. Kinetic analysis utilizing fluorescent resonance energy transfer (FRET), that is only observed upon the heterodimeric ECFP-EYFP conjugation, revealed that the amino acid replacement contributed to the acceleration of cross-linking reactions by increasing catalytic turnover (kcat), rather than substrate binding affinity (Km). Finally, using a ribonuclease S-protein, the manipulation of enzymatic protein cross-linking based on specific S-peptide:S-protein interactions was explored. Since newly designed Lys- and Gln-tags retained binding affinities to the S-protein, the heterodimerization was perfectly restrained by wrapping them with the S-protein. The results suggest the possibility of limited protein conjugation by tuning steric hindrance against the MTG. Tailoring enzymatic posttranslational modifications with either engineering peptidyl substrates or by taking specific peptide-protein interactions into consideration may facilitate the development of a new sequential protein conjugation method for the preparation of multifunctional protein.. |
| 496. |
T Takazawa, N Kamiya, H Ueda, T Nagamune, Enzymatic labeling of a single chain variable fragment of an antibody with alkaline phosphatase by mircobial transglutaminase, BIOTECHNOLOGY AND BIOENGINEERING, 10.1002/bit.20019, 86, 4, 399-404, 2004.05, Functional cross-linking of a single chain Fv fragment of anti-hen egg-white lysozyme antibody (scFv) and alkaline phosphatase (AP) was explored using microbial transglutaminase (MTG) from Streptomyces mobaraensis. A specific pepticlyl linker for MTG was genetically fused to the N-terminus of each protein and the resultant proteins were obtained separately by bacterial expression. The recombinant peptide-tagged scFv and AP were site-specifically cross-linked by MTG through the extra peptidyl linkers in vitro, which mainly yielded the heterodimer (i.e., scFv-AP conjugate). The enzymatic cross-linking reaction had little influence on either the antigen-binding ability of the scFv moiety or the enzymatic activity of the AP moiety of the conjugate, allowing use within an enzyme-linked immunosorbent assay. The results obtained suggest that the enzymatic approach with MTG facilitates the posttranslational construction of functional fusion proteins. (C) 2004 Wiley Periodicals, Inc.. |
| 497. |
T Tanaka, N Kamiya, T Nagamune, Peptidyl linkers for protein heterodimerization catalyzed by microbial transglutaminase, BIOCONJUGATE CHEMISTRY, 10.1021/bc034209o, 15, 3, 491-497, 2004.05, Specific peptidyl linkers that result in the heterodimerization of functional proteins, which is catalyzed by microbial transglutaminase from Streptomyces mobaraensis (MTG), were generated based on a ribonuclease S-peptide using site-directed mutagenesis. The peptidyl linkers designated as Lys-tag and Gln-tag were designed to possess sole reactive Lys or Gln residue that was amenable for selective Lys-Gln cross-linkage of different proteins. Green fluorescent protein variants, ECFP and EYFP, were employed as model proteins, and those Lys- and Gln-tags were fused to the N-termini of ECFP and EYFP, respectively. As a result, we succeeded in solely obtaining the ECFP-EYFP heterodimer without forming multiply cross-linked byproducts. It was found that the reactivity of peptidyl linkers varied according to the type of amino acid to be replaced. Peptidyl linkers with a basic amino acid (Arg) exhibited the highest reactivity in the cross-linking reaction, suggesting the cationic residue substrate preference of MTG. Kinetic analysis utilizing fluorescent resonance energy transfer (FRET), that is only observed upon the heterodimeric ECFP-EYFP conjugation, revealed that the amino acid replacement contributed to the acceleration of cross-linking reactions by increasing catalytic turnover (k(cat))rather than substrate binding affinity (K-m). Finally, using a ribonuclease S-protein, the manipulation of enzymatic protein cross-linking based on specific S-peptide:S-protein interactions was explored. Since newly designed Lys- and Gln-tags retained binding affinities to the S-protein, the heterodimerization was perfectly restrained by wrapping them with the S-protein. The results suggest the possibility of limited protein conjugation by tuning steric hindrance against the MTG. Tailoring enzymatic posttranslational modifications with either engineering peptidyl substrates or by taking specific peptide-protein interactions into consideration may facilitate the development of a new sequential protein conjugation method for the preparation of multifunctional protein.. |
| 498. |
Tatsuo Maruyama, Hiroshi Yamamura, Takahiro Kotani, Noriho Kamiya, Masahiro Goto, Poly(ethylene glycol)-lipase complexes that are highly active and enantioselective in ionic liquids, Organic and Biomolecular Chemistry, 10.1039/b401012d, 2, 8, 1239-1244, 2004.04, Lipase-catalyzed alcoholysis between vinyl acetate and 2-phenyl-1-propanol was investigated in dialkylimidazolium-based ionic liquids. Although native lipase powder exhibited very low activity in an ionic liquid, forming a polyethylene glycol) (PEG)-lipase complex improved the lipase activity in the ionic liquid. The activity of the PEG-lipase complex was higher in ionic liquids than in common organic solvents (n-hexane, isooctane and dimethylsulfoxide). Fluorescence measurements using 4-aminophthalimide revealed that the ionic liquids were more hydrophilic than the organic solvents used for non-aqueous enzymology. A kinetic study of lipase-catalyzed alcoholysis in an ionic liquid ([Bmim][PF6]) revealed that the Michaelis constant (Km) for 2-phenyl-1-propanol in the ionic liquid was half that in n-hexane, suggesting that the ionic liquid stabilized the enzyme-substrate complex. Finally, we carried out enantioselective alcoholysis of 1-phenylethanol in ionic liquids employing the PEG-lipase complex, and obtained high enantioselectivity, comparable to that in n-hexane.. |
| 499. |
Tatsuo Maruyama, Jun Ichi Uchida, Tomohiro Ohkawa, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Solid-phase peptide synthesis in a microfluidic device, kagaku kogaku ronbunshu, 10.1252/kakoronbunshu.30.180, 30, 2, 180-182, 2004.03, The solid-phase synthesis of oligopeptide was achieved for the first time in a microfluidic device. We fabricated a microchannel (width 300 μm, depth 160 μm) with a bead trap on a glass plate. Resin beads were loaded in the bead trap for the solid-phase peptide synthesis, and the synthesis of pentapeptide (Met-enkephalin) was conducted in the microchannel. The yield of the synthesized pentapeptide was confirmed to be 20% by MALDI-TOF MS and HPLC analyses.. |
| 500. |
マイクロ流体デバイスを用いたオリゴペプチドの固相合成. |
| 501. |
Tatsuo Maruyama, Takahiro Kotani, Hiroshi Yamamura, Noriho Kamiya, Masahiro Goto, Poly(ethylene glycol)-lipase complexes catalytically active in fluorous solvents, Organic and Biomolecular Chemistry, 10.1039/b312212c, 2, 4, 524-527, 2004.02, Lipase-catalyzed alcoholysis between vinyl cinnamate and benzyl alcohol in fluorous solvents was investigated. This is the first report of a lipase-catalyzed reaction in a fluorous solvent. Forming the poly(ethylene glycol) (PEG)-lipase PL complex enhanced lipase activity over 16-fold over that of native lipase powder. The PEG-lipase PL complex exhibited markedly higher alcoholysis activities in fluorous solvents than in conventional organic solvents such as isooctane and n-hexane. The optimum reaction temperature for FC-77 (perfluorooctane) was 55°C and the optimum pH for the preparation of the PEG-lipase complex was 9.0; similar to the conditions for lipase PL-catalyzed reaction in aqueous solution. The alcoholysis reaction in fluorous solvent requires the addition of a FC77-miscible organic solvent (isooctane) in order to dissolve non-fluorinated substrates. Lipase activity in the fluorous solvent was significantly influenced by the volume ratio of isooctane in the reaction medium. Vinyl cinnamate inhibition of the lipase-catalyzed reaction occurred at a much lower concentration in the fluorous solvent than in isooctane. These results can be explained by the localization of substrates around lipase molecules, induced by adsorption of the substrates to the PEG layer of the PEG-lipase complex.. |
| 502. |
T Maruyama, T Kotani, H Yamamura, N Kamiya, M Goto, Poly(ethylene glycol)-lipase complexes catalytically active in fluorous solvents, ORGANIC & BIOMOLECULAR CHEMISTRY, 10.1039/b312212c, 2, 4, 524-527, 2004.02, Lipase-catalyzed alcoholysis between vinyl cinnamate and benzyl alcohol in fluorous solvents was investigated. This is the first report of a lipase-catalyzed reaction in a fluorous solvent. Forming the poly(ethylene glycol) (PEG)-lipase PL complex enhanced lipase activity over 16-fold over that of native lipase powder. The PEG-lipase PL complex exhibited markedly higher alcoholysis activities in fluorous solvents than in conventional organic solvents such as isooctane and n-hexane. The optimum reaction temperature for FC-77 (perfluorcooctane) was 55degreesC and the optimum pH for the preparation of the PEG-lipase complex was 9.0; similar to the conditions for lipase PL-catalyzed reaction in aqueous solution. The alcoholysis reaction in fluorous solvent requires the addition of a FC77-miscible organic solvent (isooctane) in order to dissolve non-fluorinated substrates. Lipase activity in the fluorous solvent was significantly influenced by the volume ratio of isooctane in the reaction medium. Vinyl cinnamate inhibition of the lipase-catalyzed reaction occurred at a much lower concentration in the fluorous solvent than in isooctane. These results can be explained by the localization of substrates around lipase molecules, induced by adsorption of the substrates to the PEG layer of the PEG-lipase complex.. |
| 503. |
H.Ichinose, J.Michizoe, T.Maruyama, N.Kamiya, M.Goto, Electron transfer reaction and functionalization of P450cam system in reverse micelles, Langmuir, 10.1021/la049752n, 20, 13, 5564-5568, vol.20, 5564-5568 (2004), 2004.01. |
| 504. |
Tatsuo Maruyama, Jun Ichi Uchida, Tomohiro Ohkawa, Toru Futami, Koji Katayama, Kei Ichiro Nishizawa, Ken Ichiro Sotowa, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Erratum
Enzymatic degradation of p-chlorophenol in a two-phase flow microchannel system (Lab on a chip - Miniaturisation for Chemistry, Biology & Bioengineering (2003) 3 (308) (DOI: 10.1039/b309982b)), Lab on a Chip, 4, 2, 2004.01. |
| 505. |
Jo Tominaga, Junji Michizoe, Noriho Kamiya, Hirofumi Ichinose, Tatsuo Maruyama, Masahiro Goto, Factors affecting the oxidative activity of laccase towards biphenyl derivatives in homogeneous aqueous-organic systems, Journal of Bioscience and Bioengineering, 10.1016/S1389-1723(04)70236-4, 98, 1, 14-19, 2004.01, Catalytic oxidation of biphenyl derivatives was investigated using laccase in a homogeneous aqueous-organic system. A thermostable laccase from Trametes sp. showed the highest catalytic activity for the oxidation of 4-hydroxybiphenyl (4-HB) at a reaction temperature of 60°C when dimethylsulfoxide (DMSO) was employed as a co-solvent. Furthermore, the catalytic performance was successfully enhanced by the incorporation of a laccase mediator system (LMS) into the aqueous-DMSO media. The catalytic performance strongly depended on the type of mediator, and the highest activity was observed with 2,2′-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) as mediator, suggesting the importance of the selection of a suitable mediator. It was verified that this mediator system is applicable to the oxidation of several biphenyl derivatives with hydroxyl groups.. |
| 506. |
Hidehiko Hirakawa, Noriho Kamiya, Yutaka Kawarabayashi, Teruyuki Nagamune, Properties of an alcohol dehydrogenase from the hyperthermophilic archaeon Aeropyrum pernix K1, Journal of Bioscience and Bioengineering, 10.1263/jbb.97.202, 97, 3, 202-206, 2004.01, A NAD+-dependent medium-chain alcohol dehydrogenase from the hyperthermophilic archaeon Aeropyrum pernix K1 was expressed in Escherichia coli and purified. The recombinant enzyme was a homotetramer of molecular mass 1.6×102 kDa. The optimum pH for the oxidative reaction was around 10.5 and that for the reductive reaction was around 8.0. The enzyme had a broad substrate specificity including aliphatic and aromatic alcohols, aliphatic and aromatic ketones, and benzylaldehyde. This enzyme produced (S)-alcohols from the corresponding ketones. The enzyme was thermophilic and the catalytic activity increased up to 95°C. It maintained 24% of the original catalytic activity after incubation for 30 min at 98°C, indicating that this enzyme is highly thermostable.. |
| 507. |
Shuji Takeda, Noriho Kamiya, Teruyuki Nagamune, Rational design of a protein-based molecular device consisting of blue fluorescent protein and zinc protoporphyrin IX incorporated into a cytochrome b562 scaffold, Biotechnology Letters, 10.1023/B:BILE.0000012889.24412.ca, 26, 2, 121-125, 2004.01, To make a single molecular photo-device, it is essential to control the exact orientation of two types of proteins. We made a chimeric protein in which cytochrome b562 was linked to the N-terminus of enhanced green fluorescent protein, cytb562-EGFP. Within cytb562-EGFP, the excitation energy of EGFP was transferred to the cytochrome b562 cofactor fixed proximally to EGFP. Cytb562-EGFP was engineered so that iron protoporphyrin IX was substituted by zinc protoporphyrin IX to make it a suitable cofactor for photo-induced electron transfer. The photosensitizer pigment was optimized and the EGFP was replaced by a blue fluorescent mutant that gave 15% higher energy transfer efficiency.. |
| 508. |
Erratum: Enzymatic degradation of p-chlorophenol in a two-phase flow microchannel system (Lab on a chip - Miniaturisation for Chemistry, Biology & Bioengineering (2003) 3 (308) (DOI: 10.1039/b309982b)). |
| 509. |
S Takeda, N Kamiya, T Nagamune, Rational design of a protein-based molecular device consisting of blue fluorescent protein and zinc protoporphyrin IX incorporated into a cytochrome b(562) scaffold, BIOTECHNOLOGY LETTERS, 10.1023/B:BILE.0000012889.24412.ca, 26, 2, 121-125, 2004.01, To make a single molecular photo-device, it is essential to control the exact orientation of two types of proteins. We made a chimeric protein in which cytochrome b(562) was linked to the N-terminus of enhanced green fluorescent protein, cytb(562)-EGFP. Within cytb(562)-EGFP, the excitation energy of EGFP was transferred to the cytochrome b562 cofactor fixed proximally to EGFP. Cytb(562)-EGFP was engineered so that iron protoporphyrin IX was substituted by zinc protoporphyrin IX to make it a suitable cofactor for photo-induced electron transfer. The photosensitizer pigment was optimized and the EGFP was replaced by a blue fluorescent mutant that gave 15% higher energy transfer efficiency.. |
| 510. |
Tatsuo Maruyama, Jun Ichi Uchida, Tomohiro Ohkawa, Toru Futami, Koji Katayama, Kei Ichiro Nishizawa, Ken Ichiro Sotowa, Fukiko Kubota, Noriho Kamiya, Masahiro Goto, Enzymatic degradation of p-chlorophenol in a two-phase flow microchannel system, Lab on a Chip, 10.1039/b309982b, 3, 4, 308-312, 2003.11, Enzymatic degradation of p-chlorophenol was carried out in a two-phase flow in a microchannel (100 μm width, 25 μm depth) fabricated on a glass plate (70 mm × 38 mm). This is the first report on the enzymatic reaction in a two-phase flow on a microfluidic device. The surface of the microchannel was partially modified with octadecylsilane groups to be hydrophobic, thus allowing clear phase separation at the end-junction of the microchannel. The enzyme (laccase), which is surface active, was solubilized in a succinic aqueous buffer and the substrate (p-chlorophenol) was in isooctane. The degradation of p-chlorophenol occurred mainly at the aqueous-organic interface in the microchannel. We investigated the effects of flow velocity and microchannel shape on the enzymatic degradation of p-chlorophenol. Assuming that diffusion of the substrate (p-chlorophenol) is the rate-limiting step in the enzymatic degradation of p-chlorophenol in the microchannel, we proposed a simple theoretical model for the degradation in the microchannel. The calculated degradation values agreed well with the experimental data.. |
| 511. |
H Hirakawa, N Kamiya, T Yata, T Nagamune, Regioselective reduction of a steroid in a reversed micellar system with enzymatic NADH-regeneration, BIOCHEMICAL ENGINEERING JOURNAL, 10.1016/S1369-703X(03)00019-6, 16, 1, 35-40, 2003.10, The regioselective reduction of androstandione to androsterone by 3alpha-hydroxysteroid dehydrogenase (HSDH) from Pseudomonas testosteroni was coupled to a cofactor (NADH) regeneration system with the oxidation of ethanol by yeast alcohol dehydrogenase (YADH) in sodium dioctyl sulfosuccinate (AOT)/isooctane reversed micelles. Each reaction was dominated by the system's water content (W-0 = [H2O]/[AOT]). The catalytic activity of YADH increased monotonically with increasing W-0, whereas HSDH showed bell-shaped dependency on W-0. Using a reversed micellar system increased stability of both enzymes in comparison with an aqueous system and prolonged NADH-regeneration; namely, a seven-fold increase in the total turnover number of NADH was achieved. These results suggest that the reversed micellar system is a promising choice for conjugate enzymatic reactions with dehydrogenases in a nonaqueous media. (C) 2003 Elsevier Science B.V.. All rights reserved.. |
| 512. |
ナノ集合体を利用した有機溶媒中におけるマンガンペルオキシダーゼの機能発現と環境汚染物質の分解. |
| 513. |
N Kamiya, T Takazawa, T Tanaka, H Ueda, T Nagamune, Site-specific cross-linking of functional proteins by transglutamination, ENZYME AND MICROBIAL TECHNOLOGY, 10.1016/S0141-0229(03)00154-6, 33, 4, 492-496, 2003.09, Microbial transglutaminase (MTG), an enzyme that works in posttranslational modification of proteins, has been applied to in vitro preparation of a bi-functional fusion protein from separately obtained recombinant proteins. A quite simple strategy in which a specific peptidyl linker recognizable by MTG is fused to the N-terminus of proteins of interest has been verified in the preparation of a bi-functional fusion protein using an antibody variable domain (single chain Fv of anti-hen-egg white lysozyme antibody, scFv) and a fluorescent protein (enhanced yellow fluorescent protein, EYFP). The resultant peptidyl linker-fused proteins were readily cross-linked by MTG to only give the heterodimer (i.e. scFv-EYFP fusion protein), suggesting that the reaction proceeded in highly specific manner. As the fusion protein exhibited sufficient bi-functionality in fluorescence immunoassay (FIA), this work shows for the first time a successful enzymatic preparation of a bi-functional fusion protein through recombinantly incorporated specific peptidyl linkers into target proteins. (C) 2003 Elsevier Inc. All rights reserved.. |
| 514. |
Tamotsu Zako, Keiichi Ayabe, Takahide Aburatani, Noriho Kamiya, Atsushi Kitayama, Hiroshi Ueda, Teruyuki Nagamune, Luminescent and substrate binding activities of firefly luciferase N-terminal domain, Biochimica et Biophysica Acta - Proteins and Proteomics, 10.1016/S1570-9639(03)00179-1, 1649, 2, 183-189, 2003.07, Firefly luciferase catalyzes highly efficient emission of light from the substrates luciferin, Mg-ATP, and oxygen. A number of amino acid residues are identified to be important for the luminescent activity, and almost all the key residues are thought to be located in the N-terminal domain (1-437), except one in the C-terminal domain, Lys529, which is thought to be critical for efficient substrate orientation. Here we show that the purified N-terminal domain still binds to the substrates luciferin and ATP with reduced affinity, and retains luminescent activity of up to 0.03% of the wild-type enzyme (WT), indicating that all the essential residues for the activity are located in the N-terminal domain. Also found is low luminescence enhancement by coenzyme A (CoA), which implies a lower product inhibition than in the WT enzyme. These findings have interesting implications for the light emission reaction mechanism of the enzyme, such as reaction intermediates, product inhibition, and the role of the C-terminal domain.. |
| 515. |
Satoshi Yamaguchi, Teruhisa Mannen, Tamotsu Zako, Noriho Kamiya, Teruyuki Nagamune, Measuring adsorption of a hydrophobic probe with a surface plasmon resonance sensor to monitor conformational changes in immobilized proteins, Biotechnology Progress, 10.1021/bp034015n, 19, 4, 1348-1354, 2003.07, Conformational changes of proteins immobilized on solid matrices were observed by measuring the adsorption of Triton X-100 (TX), a nonionic detergent, as a hydrophobic probe with BIACORE, a biosensor that utilizes the phenomenon of surface plasmon resonance (SPR). Two kinds of proteins, α-glucosidase and lysozyme, were covalently attached to dextran matrices on the sensor surface in the flow cell and then exposed to various concentrations of TX solution. We measured SPR signal changes derived from adsorption of TX to the immobilized proteins and calculated the monolayer adsorption capacity using the Brunauer-Emmett-Teller (BET) equation. The results demonstrated that monolayer adsorption capacity is proportional to the amount of immobilized proteins. Further, the unfolding process of immobilized proteins on the sensor surface induced by guanidine hydrochloride was investigated by monitoring SPR signal increases due to the adsorption of TX to the exposed hydrophobic region of the protein. Results strongly suggested that the increase in the SPR signal reflected the formation of the agglutinative unfolded state. We expect our measuring method using the SPR sensor and TX adsorption will be a novel tool to provide conformational information regarding various proteins on solid matrices.. |
| 516. |
T Zako, K Ayabe, T Aburatani, N Kamiya, A Kitayama, H Ueda, T Nagamune, Luminescent and substrate binding activities of firefly luciferase N-terminal domain, BIOCHIMICA ET BIOPHYSICA ACTA-PROTEINS AND PROTEOMICS, 10.1016/S1570-9639(03)00179-1, 1649, 2, 183-189, 2003.07, Firefly luciferase catalyzes highly efficient emission of light from the substrates luciferin, Mg-ATP, and oxygen. A number of amino acid residues are identified to be important for the luminescent activity, and almost all the key residues are thought to be located in the N-terminal 529 domain (1-437), except one in the C-terminal domain, Lys(529), which is thought to be critical for efficient substrate orientation. Here we show that the purified N-terminal domain still binds to the substrates luciferin and ATP with reduced affinity, and retains luminescent activity of up to 0.03% of the wild-type enzyme (WT), indicating that all the essential residues for the activity are located in the N-terminal domain. Also found is low luminescence enhancement by coenzyme A (CoA), which implies a lower product inhibition than in the WT enzyme. These findings have interesting implications for the light emission reaction mechanism of the enzyme, such as reaction intermediates, product inhibition, and the role of the C-tenninal domain. (C) 2003 Elsevier B.V All rights reserved.. |
| 517. |
S Yamaguchi, T Mannen, T Zako, N Kamiya, T Nagamune, Measuring adsorption of a hydrophobic probe with a surface plasmon resonance sensor to monitor conformational changes in immobilized proteins, BIOTECHNOLOGY PROGRESS, 10.1021/bp034015n, 19, 4, 1348-1354, 2003.07, Conformational changes of proteins immobilized on solid matrices were observed by measuring the adsorption of Triton X-100 (TX), a nonionic detergent, as a hydrophobic probe with BIACORE, a biosensor that utilizes the phenomenon of surface plasmon resonance (SPR). Two kinds of proteins, a-glucosidase and lysozyme, were covalently attached to dextran matrices on the sensor surface in the flow cell and then exposed to various concentrations of TX solution. We measured SPR signal changes derived from adsorption of TX to the immobilized proteins and calculated the monolayer adsorption capacity using the Brunauer-Emmett-Teller (BET) equation. The results demonstrated that monolayer adsorption capacity is proportional to the amount of immobilized proteins. Further, the unfolding process of immobilized proteins on the sensor surface induced by guanidine hydrochloride was investigated by monitoring SPR signal increases due to the adsorption of TX to the exposed hydrophobic region of the protein. Results strongly suggested that the increase in the SPR signal reflected the formation of the agglutinative unfolded state. We expect our measuring method using the SPR sensor and TX adsorption will be a novel tool to provide conformational information regarding various proteins on solid matrices.. |
| 518. |
Shuji Takeda, Noriho Kamiya, Teruyuki Nagamune, A novel protein-based heme sensor consisting of green fluorescent protein and apocytochrome b562, Analytical Biochemistry, 10.1016/S0003-2697(03)00096-4, 317, 1, 116-119, 2003.06. |
| 519. |
Noriho Kamiya, Alexander M. Klibanov, Controling the rate of protein release from polyelectrolyte complexes, Biotechnology and Bioengineering, 10.1002/bit.10606, 82, 5, 590-594, 2003.06, Extended protein release from readily prepared, water-insoluble complexes with oppositely charged polyions is explored. Using hen egg-white lysozyme as a model, its sustained release from such complexes with a number of polyanions under physiological conditions has been demonstrated and rationalized. The rate of release varies orders of magnitude and is controlled by the nature of the polyanion (decreasing upon increase in its linear charge density, length, and hydrophobicity) and the complex particle size (the larger the particles, the slower the release).. |
| 520. |
S Takeda, N Kamiya, T Nagamune, A novel protein-based heme sensor consisting of green fluorescent protein and apocytochrome b(562), ANALYTICAL BIOCHEMISTRY, 10.1016/S-0003-2697(03)00096-4, 317, 1, 116-119, 2003.06. |
| 521. |
N Kamiya, AM Klibanov, Controling the rate of protein release from polyelectrolyte complexes, BIOTECHNOLOGY AND BIOENGINEERING, 10.1002/bit.10606, 82, 5, 590-594, 2003.06, Extended protein release from readily prepared, water-insoluble complexes with oppositely charged polyions is explored. Using hen egg-white lysozyme as a model, its sustained release from such complexes with a number of polyanions under physiological conditions has been demonstrated and rationalized. The rate of release varies orders of magnitude and is controlled by the nature of the polyanion (decreasing upon increase in its linear charge density, length, and hydrophobicity) and the complex particle size (the larger the particles, the slower the release). (C) 2003 Wiley Periodicals, Inc.. |
| 522. |
Bumhwan Lee, Noriho Kamiya, Shinjiro Machida, Yutaka Yamagata, Kazuyuki Horie, Teruyuki Nagamune, Fabrication of a protein film by electrospray deposition method and investigation of photochemical properties by persistent spectral hole burning, Biomaterials, 10.1016/S0142-9612(02)00637-3, 24, 12, 2045-2051, 2003.05, Transparent protein film of iron-free cytochrome c (Cyt. c) was successfully manufactured by electrospray deposition (ES) method and the properties of the protein film were investigated. Excursion temperature dependence of spectral hole profiles in the photochemical hole burning for the iron-free Cyt. c in protein film and in glassy solution was investigated and compared with that of iron-free porphyrin embedded in a synthetic polymer film to clarify photochemical properties of electrospray deposited protein film. The spectral holes of iron-free porphyrin were thermally more stable in Cyt. c than in polymer, indicating compact packing of the chromophore in the protein. The ES-deposited iron-free Cyt. c in protein film showed less stable spectral hole than that in glassy solution. This difference is attributable to the electrospray deposition process of the protein involving ionization and to subsequent cross-linking of protein molecules.. |
| 523. |
B Lee, N Kamiya, S Machida, Y Yamagata, K Horie, T Nagamune, Fabrication of a protein film by electrospray deposition method and investigation of photochemical properties by persistent spectral hole burning, BIOMATERIALS, 10.1016/S0142-9612(02)00637-3, 24, 12, 2045-2051, 2003.05, Transparent protein film of iron-free cytochrome c (Cyt. c) was successfully manufactured by electrospray deposition (ES) method and the properties of the protein film were investigated. Excursion temperature dependence of spectral hole profiles in the photochemical hole burning for the iron-free Cyt. c in protein film and in glassy solution was investigated and compared with that of iron-free porphyrin embedded in a synthetic polymer film to clarify photochemical properties of electrospray deposited protein film. The spectral holes of iron-free porphyrin were thermally more stable in Cyt. c than in polymer, indicating compact packing of the chromophore in the protein. The ES-deposited iron-free Cyt. c in protein film showed less stable spectral hole than that in glassy solution. This difference is attributable to the electrospray deposition process of the protein involving ionization and to subsequent cross-linking of protein molecules. (C) 2003 Elsevier Science Ltd. All rights reserved.. |
| 524. |
E Miyako, T Maruyama, N Kamiya, M Goto, Use of ionic liquids in a lipase-facilitated supported liquid membrane, BIOTECHNOLOGY LETTERS, 10.1023/A:1023536922749, 25, 10, 805-808, 2003.05, A lipase-facilitated transport of 4-phenoxybutyric acid, 3-phenoxypropionic acid, 2-phenylpropionic acid, 2-phenoxybutyric acid, mandelic acid and 2-amino-2-phenylbutyric acid was carried out using a supported liquid membrane based on room temperature ionic liquids. There were marked differences in the permeate fluxes of various organic acids due to the substrate specificity of the lipases. The maximum permeate flux (44 x 10(-2) mmol cm(-2) x h) was obtained using 4-phenoxybutyric acid as the substrate and 1-n-butyl-3-methylimidazolium hexafluorophosphate as the liquid membrane phase.. |
| 525. |
Eiichi Toorisaka, Yuko Kokazu, Noriho Kamiya, Masahiro Goto, Leakage mechanism of irinotecan from water-in-oil-in-water (W/O/W) multiple emulsions, kagaku kogaku ronbunshu, 10.1252/kakoronbunshu.29.294, 29, 2, 294-298, 2003.03, The leakage mechanism of the anticancer drug Irinotecan (CPT-11) from multiple W/O/W emulsions was investigated. The molecular structure of surfactants used for the preparation of the multiple emulsions affected the leakage rate of CPT-11. A sugar-long alkyl chain ester was effective in suppressing the drug release from the emulsions, because the surfactant reduces the mobility of inner aqueous droplets in emulsions. The leakage of drugs was deduced to be caused by the thin lamella membrane formed between the internal and external aqueous phases. The direction of water transportation was also an important factor in the degree of the drug release.. |
| 526. |
Noriho Kamiya, Tsutomu Tanaka, Tsutomu Suzuki, Takeshi Takazawa, Shuji Takeda, Kimitsuna Watanabe, Teruyuki Nagamune, S-peptide as a potent peptidyl linker for protein cross-linking by microbial transglutaminase from Streptomyces mobaraensis, Bioconjugate Chemistry, 10.1021/bc025610y, 14, 2, 351-357, 2003.03, We have found that ribonuclease S-peptide can work as a novel peptidyl substrate in protein cross-linking reactions catalyzed by microbial transglutaminase (MTG) from Streptomyces mobaraensis. Enhanced green fluorescent protein tethered to S-peptide at its N-terminus (S-tag-EGFP) appeared to be efficiently cross-linked by MTG. As wild-type EGFP was not susceptible to cross-linking, the S-peptide moiety is likely to be responsible for the cross-linking. A site-directed mutation study assigned Gln15 in the S-peptide sequence as the sole acyl donor. Mass spectrometric analysis showed that two Lys residues (Lys5 and Lys11) in the S-peptide sequence functioned as acyl acceptors. We also succeeded in direct monitoring of the cross-linking process by virtue of fluorescence resonance energy transfer (FRET) between S-tag-EGFP and its blue fluorescent color variant (S-tag-EBFP). The protein cross-linking was tunable by either engineering S-peptide sequence or capping the S-peptide moiety with S-protein, the partner protein of S-peptide for the formation of ribonuclease A. The latter indicates that S-protein can be used as a specific inhibitor of S-peptide-directed protein cross-linking by MTG. The controllable protein cross-linking of S-peptide as a potent substrate of MTG will shed new light on biomolecule conjugation.. |
| 527. |
T Maruyama, S Noda, N Kamiya, M Goto, Ring-opening polymerization of lactones catalyzed by surfactant-coated lipases in organic solvents, JOURNAL OF CHEMICAL ENGINEERING OF JAPAN, 10.1252/jcej.36.307, 36, 3, 307-312, 2003.03, We have performed the ring-opening polymerization of lactones catalyzed by surfactant-coated lipases in organic solvents. A great improvement in the number-average molecular weight (M-n) of the produced polymer was observed compared to that of the lyophilized native lipase. The M-n vs monomer conversion profile suggests that the polymerization catalyzed by the surfactant-coated lipase is a step polymerization, whereas that catalyzed by lyophilized lipase powder is a chain polymerization. The effects of reaction temperature and the reaction solvent on the polymerization of lactones were also investigated.. |
| 528. |
N Kamiya, T Tanaka, T Suzuki, T Takazawa, S Takeda, K Watanabe, T Nagamune, S-peptide as a potent peptidyl linker for protein cross-linking by microbial transglutaminase from Streptomyces mobaraensis, BIOCONJUGATE CHEMISTRY, 10.1021/bc025610y, 14, 2, 351-357, 2003.03, We have found that ribonuclease S-peptide can work as a novel peptidyl substrate in protein cross-linking reactions catalyzed by microbial transglutaminase (MTG) from Streptomyces mobaraensis. Enhanced green fluorescent protein tethered to S-peptide at its N-terminus (S-tag-EGFP) appeared to be efficiently cross-linked by MTG. As wild-type EGFP was not susceptible to cross-linking, the S-peptide moiety is likely to be responsible for the cross-linking. A site-directed mutation study assigned Gln15 in the S-peptide sequence as the sole acyl donor. Mass spectrometric analysis showed that two Lys residues (Lys5 and Lys11) in the S-peptide sequence functioned as acyl acceptors. We also succeeded in direct monitoring of the cross-linking process by virtue of fluorescence resonance energy transfer (FRET) between S-tag-EGFP and its blue fluorescent color variant (S-tag-EBFP). The protein cross-linking was tunable by either engineering S-peptide sequence or capping the S-peptide moiety with S-protein, the partner protein of S-peptide for the formation of ribonuclease A. The latter indicates that S-protein can be used as a specific inhibitor of S-peptide-directed protein cross-linking by MTG. The controllable protein cross-linking of S-peptide as a potent substrate of MTG will shed new light on biomolecule conjugation.. |
| 529. |
E Toorisaka, H Ono, K Arimori, N Kamiya, M Goto, Hypoglycemic effect of surfactant-coated insulin solubilized in a novel solid-in-oil-in-water (S/O/W) emulsion, INTERNATIONAL JOURNAL OF PHARMACEUTICS, 10.1016/S0378-5173(02)00674-9, 252, 1-2, 271-274, 2003.02, A novel solid-in-oil-in-water (S/O/W) emulsion for oral administration of insulin has been developed using surfactant-coated insulin. The S/O/W emulsion prepared by a shirasu porous glass (SPG) membrane provided a sharp size distribution and was stable. Leakage of insulin from the S/O/W emulsions was not observed for several days. The S/O/W emulsion showed the hypoglycemic activity for a long period after oral administration to rats. (C) 2002 Elsevier Science B.V. All rights reserved.. |
| 530. |
Junji Michizoe, Yoichi Uchimura, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Control of water content by reverse micellar solutions for peroxidase catalysis in a water-immiscible organic solvent, Journal of Bioscience and Bioengineering, 10.1263/jbb.95.425, 95, 4, 425-427, 2003.01, The water content of a water-immiscible can be controlled using reverse micelles. We applied this reverse micellar system to improve the enzymatic activity of a surfactant-manganese peroxidase complex in toluene. Increasing the water content in toluene to 2 vol% using the reverse micelles resulted in the great improvement (10-fold) of the peroxidase activity.. |
| 531. |
Eijiro Miyako, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto, Transport of Organic Acids through a Supported Liquid Membrane Driven by Lipase-Catalyzed Reactions, Journal of Bioscience and Bioengineering, 10.1263/jbb.96.370, 96, 4, 370-374, 2003.01, We have developed a lipase-facilitated supported liquid membrane. Lipase-catalyzed reactions were coupled with a supported liquid membrane (SLM) to transport organic acids through the SLM. We succeeded in the rational transport of organic acids through the SLM using lipase-catalyzed reactions and observed that there were differences in the transport behavior of various organic acids due to the substrate specificity of lipase. Subsequently, various parameters, such as the alcohol concentration in the feed phase, the pH in each aqueous phase, an organic solvent in the SLM, and the kind of lipase, were investigated. We found that the optimum conditions were 65 vol% alcohol concentration, pH 6.3 in each aqueous phase, isooctane as the liquid membrane phase and Candida rugosa lipase as the esterification biocatalyst.. |
| 532. |
Masafumi Shimizu, Noriho Kamiya, Atsushi Kitayama, Teruyuki Nagamune, Self-assembly of electron transport protein using oligonucleotide hybridization, Colloids and Surfaces B: Biointerfaces, 10.1016/S0927-7765(01)00305-8, 25, 1, 69-79, 2002.03, In order to construct a supramolecular architecture composed of an electron transport protein and oligonucleotide, two complementary oligonucleotides were appended to cytochrome b562 (b562) and its cofactor heme as 'tab for sticking'. His 63 of b562, located at the opposite side of the heme crevice was replaced with Cys to generate b562-SH. A 24-mer linker oligonucleotide (LO) was successfully linked through the Cys 63 residue to produce b562-LO. Moreover, the complementary oligonucleotide to LO (cLO) was appended to the heme propionate to create heme-cLO. The apoprotein of b562-SH was immobilized on the sensor surface by disulfide exchange reaction. The heme-cLO was incorporated into the immobilized apo b562-SH by the heme reconstitution, and then b562-LO was integrated on that surface by hybridization. All the self-assembly processes of these molecules by non-covalent bonding interactions were confirmed with the surface plasmon resonance biosensor. The utilization of DNA hybridization and/or apoprotein-cofactor interaction may allow a new strategy to construct a molecular electronic device.. |
| 533. |
Noriho Kamiya, Teruyuki Nagamune, Effect of water activity control on the catalytic performance of surfactant - Arthromyces ramosus peroxidase complex in toluene, Biochemical Engineering Journal, 10.1016/S1369-703X(01)00162-0, 10, 1, 55-59, 2002.02, Arthromyces ramosus peroxidase (ARP) was successfully modified with a synthetic surfactant for one-electron oxidation reaction of a hydrophobic substrate in toluene. Although UV-visible absorption spectrum of surfactant-ARP complex in toluene showed slight red shift of Soret band compared to that in water, the complex can catalyze oxidation reaction of o-phenylenediamine (o-PDA) with hydrogen peroxide. It appeared that thermodynamic water activity in the reaction system has dominant effect on either the catalytic activity or the stability in the catalytic cycle. Steady-state kinetics under the optimal condition revealed that the specific constant (kcat/Km) of ARP complex for o-PDA was 2 orders of magnitude lower than that in aqueous media, while only 13-fold lower for hydrogen peroxide. The reduction of catalytic activity caused by altering the reaction media from water to toluene was found to be mainly due to the low specific constant of ARP complex for o-PDA rather than hydrogen peroxide.. |
| 534. |
N Kamiya, T Nagamune, Effect of water activity control on the catalytic performance of surfactant - Arthromyces ramosus peroxidase complex in toluene, BIOCHEMICAL ENGINEERING JOURNAL, 10.1016/S1369-703X(01)00162-0, 10, 1, 55-59, 2002.02, Arthromyces ramosus peroxidase (ARP) was successfully modified with a synthetic surfactant for one-electron oxidation reaction of a hydrophobic substrate in toluene. Although UV-visible absorption spectrum of surfactant-ARP complex in toluene showed slight red shift of Soret band compared to that in water, the complex can catalyze oxidation reaction of o-phenylenediamine (o-PDA) with hydrogen peroxide. It appeared that thermodynamic water activity in the reaction system has dominant effect on either the catalytic activity or the stability in the catalytic cycle. Steady-state kinetics under the optimal condition revealed that the specific constant (k(cat)/K-m) of ARP complex for o-PDA was 2 orders of magnitude lower than that in aqueous media, while only 13-fold lower for hydrogen peroxide. The reduction of catalytic activity caused by altering the reaction media from water to toluene was found to be mainly due to the low specific constant of ARP complex for o-PDA rather than hydrogen peroxide. (C) 2002 Elsevier Science B.V. All rights reserved.. |
| 535. |
Shuji Takeda, Noriho Kamiya, Ryoichi Arai, Teruyuki Nagamune, Design of an artificial light-harvesting unit by protein engineering
Cytochrome b562-green fluorescent protein chimera, Biochemical and Biophysical Research Communications, 10.1006/bbrc.2001.5966, 289, 1, 299-304, 2001.11, We have generated a novel model protein for an artificial light-harvesting complex composed of two proteins, cytochrome b562 (cytb562) and enhanced green fluorescent protein (EGFP), in which two chromophores are fixed in each protein matrix. Cytb562 was appended to the N-terminus of EGFP via a Gly-Ser linker and the resultant fusion protein was successfully expressed in Escherichia coli as a mixture of the apo- and the holo-forms as to the cytb562 moiety. The fluorescence of EGFP was substantially quenched when the apo-form was reconstituted with hemin. Based on the fluorescence lifetime measurements, it appeared that light energy entrapped by EGFP is transferred to the heme of cytb562 by resonance energy transfer (energy transfer yield: 65%). Spatial organization of two chromophores using small and stable protein matrices will be promising toward the construction of an artificial light-harvesting complex by protein engineering.. |
| 536. |
S Takeda, N Kamiya, R Arai, T Nagamune, Design of an artificial light-harvesting unit by protein engineering: Cytochrome b(562)-green fluorescent protein chimera, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 10.1006/bbrc.2001.5966, 289, 1, 299-304, 2001.11, We have generated a novel model protein for an artificial light-harvesting complex composed of two proteins, cytochrome b(562) (cytb(562)) and enhanced green fluorescent protein (EGFP), in which two chromophores are fixed in each protein matrix. Cytbr(562) was appended to the N-terminus of EGFP via a Gly-Ser linker and the resultant fusion protein was successfully expressed in Escherichia coli as a mixture of the apo- and the holo-forms as to the cytb(562) moiety. The fluorescence of EGFP was substantially quenched when the apo-form was reconstituted with hemin. Based on the fluorescence lifetime measurements, it appeared that light energy entrapped by EGFP is transferred to the heme Of cytb(562) by resonance energy transfer (energy transfer yield: 65%). Spatial organization of two chromophores using small and stable protein matrices will be promising toward the construction of an artificial light-harvesting complex by protein engineering. (C) 2001 Academic Press.. |
| 537. |
N Kamiya, T Ogawa, T Nagamune, Enhancement of apparent thermostability of lipase from Rhizopus sp by the treatment with a microbial transglutaminase, BIOTECHNOLOGY LETTERS, 10.1023/A:1011936618341, 23, 19, 1629-1632, 2001.10, Crude lipase from Rhizopus sp. was moderately stable against heat treatment at 45 degreesC. However, after incubation for 1 h at 25 degreesC with Streptoverticillium transglutaminase (MTG), the half-life of crude lipase in the heat treatment was increased more than 10-fold compared to that of untreated one. The result can be ascribed by the MTGase-mediated crosslinking of contaminating proteins that affect the apparent thermostability of lipase in the crude sample.. |
| 538. |
逆ミセル中における補酵素再生を伴う酸化還元反応に関する基礎的研究. |
| 539. |
Noriho Kamiya, Yuko Okimoto, Zhen Ding, Hiroko Ohtomo, Masafumi Shimizu, Atsushi Kitayama, Hisayuki Morii, Teruyuki Nagamune, How does heme axial ligand deletion affect the structure and the function of cytochrome b562?, Protein Engineering, 10.1093/protein/14.6.415, 14, 6, 415-419, 2001.01, We have recently generated a new mutant of cytochrome b562 (cytb562) in which Met7, one of the axial heme ligands, is replaced by Ala (M7A cytb562). The M7A cytb562 can bind heme and the UV-visible absorption spectrum is of a typical high-spin ferric heme. To investigate the effect of the lack of Met7 ligation on the structural integrity of cytb562, thermal transition analyses of M7A cytb562 were conducted. From the thermodynamic parameters obtained, it is concluded that the folding of M7A cytb562 is comparable to the apoprotein despite the presence of heme. On the other hand, exogenous ligands such as cyanide and azide ions are readily bound to the heme iron, indicating that the axial coordination site is available for substrate binding. The peroxidase activity of this mutant is thus examined to evaluate new enzymatic function at this site and M7A cytb562 was found to catalyze an oxidation reaction of aromatic substrates with hydrogen peroxide. These observations demonstrate that the Met7/His102 bis-ligation to the heme iron is crucial for the stable folding of cytb562, whereas the functional conversion of cytb562 is successfully achieved by the loose folding together with the open coordination site.. |
| 540. |
R. Arai, H. Ueda, A. Kitayama, N. Kamiya, T. Nagamune, Design of the linkers which effectively separate domains of a bifunctional fusion protein, Protein Engineering, 10.1093/protein/14.8.529, 14, 8, 529-532, 2001, With the aim of separating the domains of a bifunctional fusion protein, the ability of several lengths of helix-forming peptides to separate two weakly interacting β-can domains was compared with that of flexible linkers or of a three α-helices bundle domain. We introduced helix-forming peptide linkers A(EAAAK)nA (n=2-5) between two green fluorescent protein variants, EBFP and EGFP, and investigated their spectral properties. The fluorescence resonance energy transfer from EBFP to EGFP decreased as the length of the linkers increased. The circular dichroism spectra analysis suggested that the linkers form an α-helix and the α-helical contents increased as the length of the linkers increased. The results clearly suggested the ability of the helical linkers to control the distance and reduce the interference between the domains. This 'linker engineering' may open a way to the rational design of linkers which maximize the multiple functions of fusion proteins or de novo multi-domain proteins.. |
| 541. |
Noriho Kamiya, Takashi Ogawa, Teruyuki Nagamune, Enhancement of apparent thermostability of lipase from Rhizopus sp. by the treatment with a microbial transglutaminase, Biotechnology Letters, 10.1023/A:1011936618341, 23, 19, 1629-1632, 2001, Crude lipase from Rhizopus sp. was moderately stable against heat treatment at 45 °C. However, after incubation for 1 h at 25 °C with Streptoverticillium transglutaminase (MTG), the half-life of crude lipase in the heat treatment was increased more than 10-fold compared to that of untreated one. The result can be ascribed by the MTGasemediated crosslinking of contaminating proteins that affect the apparent thermostability of lipase in the crude sample.. |
| 542. |
Noriho Kamiya, Masahito Inoue, Masahiro Goto, Nobuhumi Nakamura, Yoshinori Naruta, Catalytic and structural properties of surfactant-horseradish peroxidase complex in organic media, Biotechnology Progress, 10.1021/bp990125b, 16, 1, 52-58, 2000.01, A surfactant-horseradish peroxidase (HRP) complex that is catalytically active in organic media has been successfully prepared by a method utilizing water-in-oil (W/O) emulsions. To optimize conditions for preparation of the HRP complex, the effects of some key parameters in the aqueous phase of W/O emulsions were investigated. The surfactant-HRP complex prepared with a nonionic surfactant exhibited a high catalytic activity compared to those with a cationic or anionic surfactant in anhydrous benzene. At the preparation step, the pH of the aqueous solution had a prominent effect on the enzymatic activity of the HRP complex in organic media. Several kinds of salts present in the HRP complex could be employed to enhance the catalytic performance in organic media. However, anionic ions present in the preparation process appeared to lower the catalytic activity owing to the complexation with heme iron. UV-visible absorption spectra of the HRP complex in benzene, which were prepared from a KCN solution (pH 7.0) or an alkaline solution (pH 12), were comparable with those of native HRP in aqueous solution under the same conditions. Resonance Raman spectroscopic studies also revealed that no significant change in the coordination state of the heme iron occurred even after coating the enzyme with surfactant molecules, lyophilization, and solubilization in nonaqueous media.. |
| 543. |
N Kamiya, M Inoue, M Goto, N Nakamura, Y Naruta, Catalytic and structural properties of surfactant-horseradish peroxidase complex in organic media, BIOTECHNOLOGY PROGRESS, 10.1021/bp990125b, 16, 1, 52-58, 2000.01, A surfactant-horseradish peroxidase (HRP) complex that is catalytically active in organic media has been successfully prepared by a method utilizing water-in-oil (W/O) emulsions. To optimize conditions for preparation of the HRP complex, the effects of some key parameters in the aqueous phase of W/O emulsions were investigated The surfactant-HRP complex prepared with a nonionic surfactant exhibited a high catalytic activity compared to those with a cationic or anionic surfactant in anhydrous benzene. At the preparation step, the pH of the aqueous solution had a prominent effect on the enzymatic activity of the HRP complex in organic media. Several kinds of salts present in the HRP complex could be employed to enhance the catalytic performance in organic media. However, anionic ions present in the preparation pl process appeared to lower the catalytic activity owing to the complexation with heme iron. W-visible absorption spectra of the HRP complex in benzene, which were prepared from a KCN solution (pH 7.0) or an alkaline solution (pH 12), were comparable with those of native HRP in aqueous solution under the same conditions. Resonance Raman spectroscopic studies also revealed that no significant change in the coordination state of the heme iron occurred even after coating the enzyme with surfactant molecules, lyophilization, and solubilization in nonaqueous media.. |
| 544. |
Noriho Kamiya, Hideaki Kasagi, Masahito Inoue, Koichiro Kusunoki, Masahiro Goto, Enantioselective recognition mechanism of secondary alcohol by surfactant-coated lipases in nonaqueous media, Biotechnology and Bioengineering, 10.1002/(SICI)1097-0290(19991020)65:23.0.CO;2-U, 65, 2, 227-232, 1999.10, The enantioselective recognition mechanism of secondary alcohol by lipases originated from Candida rugosa and Pseudomonas cepacia was elucidated on the basis of the kinetic study of the esterification of alcohol with lauric acid in isooctane. To obtain inherent kinetic parameters, we utilized a surfactant-coated lipase whose conformation is considered to be an 'open' form in a homogeneous organic solvent. Based on the experimental results, the enantioselectivity of lipases was found to be derived from the difference in the V(max) values between the two enantiomers. The same result was observed when lipases of different origin and substrates with different molecular structures were applied.. |
| 545. |
N Kamiya, H Kasagi, M Inoue, K Kusunoki, M Goto, Enantioselective recognition mechanism of secondary alcohol by surfactant-coated lipases in nonaqueous media, BIOTECHNOLOGY AND BIOENGINEERING, 10.1002/(SICI)1097-0290(19991020)65:23.0.CO;2-U, 65, 2, 227-232, 1999.10, The enantioselective recognition mechanism of secondary alcohol by lipases originated from Candida rugosa and Pseudomonas cepacia was elucidated on the basis of the kinetic study of the esterification of alcohol with lauric acid in isooctane. To obtain inherent kinetic parameters, we utilized a surfactant-coated lipase whose conformation is considered to be an "open" form in a homogeneous organic solvent. Based on the experimental results, the enantioselectivity of lipases was found to be derived from the difference in the V-max values between the two enantiomers. The same result was observed when lipases of different origin and substrates with different molecular structures were applied. (C) 1999 John Wiley & Sons, Inc.. |
| 546. |
Noriho Kamiya, Masahiro Goto, Shintaro Furusaki, Surfactant-histidine-heme ternary complex as a simple artificial heme enzyme in organic media, Biotechnology and Bioengineering, 10.1002/(SICI)1097-0290(19990820)64:43.0.CO;2-H, 64, 4, 502-506, 1999.08, A surfactant-heme complex which shows peroxidase activity in organic media has been prepared by a method utilizing water-in-oil (W/O) emulsions. Both the aqueous phase pH and the type of surfactant appeared to have prominent effect on the catalytic activity of the heme complex in benzene. The catalytic efficiency of the heme complex was enhanced more than ten times by adding histidine to the aqueous phase of W/O emulsions in the preparation process. The enhancement of peroxidase activity was observed only in a nonaqueous medium due to the increase of the effective concentration of histidine as an activator. In the present study, we propose a simple preparation method for an artificial heme enzyme which works in nonaqueous media.. |
| 547. |
N Kamiya, M Goto, S Furusaki, Surfactant-histidine-heme ternary complex as a simple artificial heme enzyme in organic media, BIOTECHNOLOGY AND BIOENGINEERING, 10.1002/(SICI)1097-0290(19990820)64:43.0.CO;2-H, 64, 4, 502-506, 1999.08, A surfactant-heme complex which shows peroxidase activity in organic media has been prepared by a method utilizing water-in-oil (W/O) emulsions. Both the aqueous phase pH and the type of surfactant appeared to have prominent effect on the catalytic activity of the heme complex in benzene. The catalytic efficiency of the heme complex was enhanced more than ten times by adding histidine to the aqueous phase of W/O emulsions in the preparation process. The enhancement of peroxidase activity was observed only in a nonaqueous medium due to the increase of the effective concentration of histidine as an activator. In the present study, we propose a simple preparation method for an artificial heme enzyme which works in nonaqueous media. (C) 1999 John Wiley & Sons, Inc.. |
| 548. |
Ly Iskandar, Tsutomu Ono, Noriho Kamiya, Masahiro Goto, Fumiyuki Nakashio, Shintaro Furusaki, Catalytic properties of novel reversed micellar system on trans-esterification by α-chymotrypsin in organic media, Biochemical Engineering Journal, 10.1016/S1369-703X(98)00013-8, 2, 1, 29-33, 1998.01, Novel reversed micelles formulated by sodium dioleylphosphate (SDOLP) has been developed for enzymatic reaction in organic media. Comparison study between sodium bis-(2-ethylhexyl)-sulfosuccinate (Aerosol-OT or AOT) and SDOLP, which was carried out by investigating the reactivity of each reversed micellar system as reaction media for enzymatic transesterification reaction of N-acetyl-1-phenylalanine ethyl ester (APEE) and 1-propanol, leads to the upshot that α-chymotrypsin immobilized within SDOLP reversed micelles can persevere its high catalytic activity in isooctane. The initial reaction rate of the SDOLP reversed micellar system is almost 60 times higher than that in the AOT system. Additional kinetic results, including the Michaelis constant and the maximum reaction rate, reveal that the inner core of the SDOLP reversed micelles provides a better environment for enzymatic reaction than that of the conventional AOT reversed micelles.. |
| 549. |
Noriho Kamiya, Masahiro Goto, Preparation of surfactant-coated lipases utilizing the molecular imprinting technique, Journal of Fermentation and Bioengineering, 10.1016/S0922-338X(97)86774-8, 85, 2, 237-239, 1998.01, Modulation of the enantioselectivity of two lipases exhibiting different specificities in the esterification of 2-octanol with lauric acid was investigated by utilizing surfactant-coating and molecular imprinting techniques. The enantioselectivity of the surfactant-coated lipase from Pseudomonas cepacia (PS) was enhanced in the presence of (R)-2-octanol as a print molecule, whereas the enantioselectivity of the lipase from Candida cylindracea (AY) was hardly changed by the imprinting technique. In the case of lipase AY, however, the selectivity of the native enzyme toward the (R)-isomer was changed to a preference for the (S)-isomer as a result of being coating with surfactant molecules. The molecular imprinting effect was prominent in the case of lipase PS, its enantioselectivity in the formation of 2-octanoate in isooctane being enhanced around two-fold compared with that of the native enzyme.. |
| 550. |
N. Kamiya, S. Furusaki, M. Goto, Peroxidase activity and stability of surfactant-heme complex in nonaqueous media, Biotechnology letters, 10.1023/A:1018403518763, 19, 10, 1015-1018, 1997.11, A surfactant-heme complex was prepared from hemin using a water-in-oil emulsion with a synthetic nonionic surfactant. The heme complex was soluble in anhydrous benzene with peroxidase activity for the oxidation of o-phenylene-diamine using tert-butyl hydroperoxide as an oxidant. An absorption spectrum of the heme complex in benzene was distinct from that of free heme in an aqueous buffer solution owing to the different aggregation states in the respective solution. Moreover, the heme complex could not be decomposed in benzene even in excess of the hydroperoxide due to enhanced stability.. |
| 551. |
Shin Ya Okazaki, Noriho Kamiya, Masahiro Goto, Application of novel preparation method for surfactant-protease complexes catalytically active in organic media, Biotechnology Progress, 10.1021/bp970064m, 13, 5, 551-556, 1997.09, We have applied the newly developed method of preparing surfactant-enzyme complexes to other different enzyme sources in order to study the efficacy of this technique in the catalyzed transesterification processes. Protease from varied material sources was modified with surfactant molecules utilizing water-in-oil (W/O) emulsions, and the complex so formed proved very effective in catalyzing vinyl butyrate transesterification with benzyl alcohol in organic media. By contrast, native commercial protease and a lyophilized protease from optimum buffer solution pH hardly catalyzed the above process. Although, conventionally prepared surfactant, coated protease had shown some catalytic activity, its performance was significantly lower than that of the surfactant-protease complex assembled in the novel preparation method. The typical transesterification rates catalytically induced by surfactant-protease complexes were in the range of 7-260-fold superior relative to other catalyzed reaction systems. The preparation and reaction conditions of the surfactant-protease P (Aspergillus melleus) have been optimized through studies of the effect of aqueous pH in the W/O emulsions, the nature of the organic solvents and surfactants, and the reaction temperature. The protease complex prepared from nonionic surfactant 2C18Δ9GE at optimized aqueous pH yielded the best results in isooctane at 45 °C. This novel enzyme modification method is conceptually simple, and the resultant complexes are elucidable. It is hoped that this method will become useful, particularly in preparing enzymes that are nonaqueous media sensitive and soluble in organic solution.. |
| 552. |
S Okazaki, N Kamiya, M Goto, Application of novel preparation method for surfactant-protease complexes catalytically active in organic media, BIOTECHNOLOGY PROGRESS, 10.1021/bp970064m, 13, 5, 551-556, 1997.09, We have applied the newly developed method of preparing surfactant-enzyme complexes to other different enzyme sources in order to study the efficacy of this technique in the catalyzed transesterification processes. Protease from varied material sources was modified with surfactant molecules utilizing water-in-oil (W/O) emulsions, and the complex so formed proved very effective in catalyzing vinyl butyrate transesterification with benzyl alcohol in organic media. By contrast, native commercial protease and a lyophilized protease hom optimum buffer solution pH hat-dry catalyzed the above process. Although, conventionally prepared surfactant-coated protease had shown some catalytic activity, its performance was significantly lower than that of the surfactant-protease complex assembled in the novel preparation method. The typical transesterification rates catalytically induced by surfactant-protease complexes were in the range of 7-260-fold superior relative to other catalyzed reaction systems. The preparation and reaction conditions of the surfactant-protease P (Aspergillus melleus) have been optimized through studies of the effect of aqueous pH in the W/O emulsions, the nature of the organic solvents and surfactants, and the reaction temperature. The protease complex prepared from nonionic surfactant 2C(18)Delta(9)GE at optimized aqueous pH yielded the best results in isooctane at 45 degrees C. This novel enzyme modification method is conceptually simple, and the resultant complexes are elucidable. It is hoped that this method will become useful, particularly in preparing enzymes that are nonaqueous media sensitive and soluble in organic solution.. |
| 553. |
Shin Ya Okazaki, Noriho Kamiya, Masahiro Goto, Fumiyuki Nakashio, Enantioselective esterification of glycidol by surfactant-lipase complexes in organic media, Biotechnology letters, 10.1023/A:1018337320213, 19, 6, 541-543, 1997.07, Enantioselective esterification of glycidol has been performed with lauric acid in organic media dosed with surfactant-lipase complexes as catalysts. Lipase derived from various biomaterial sources was complexed with nonionic surfactant, dioleyl-N-D-glucono-L-glutamate, prior to use. Surfactant-lipase D (from Rhizopus delemar) complex had a higher enantioselectivity (v(R)/v(S) = 7.6) than the other lipases and the corresponding initial reaction rate was averaging 100-fold better than that of native powder lipase D in cyclohexane at 35°C.. |
| 554. |
Noriho Kamiya, Masahiro Goto, How is enzymatic selectivity of menthol esterification catalyzed by surfactant-coated lipase determined in organic media?, Biotechnology Progress, 10.1021/bp9700317, 13, 4, 488-492, 1997.07, A kinetic study of menthol esterification reaction catalyzed by surfactant-coated lipase (from Candida rugosa) has been conducted in dry isooctane. The kinetic characteristics observed in the enantioselective esterification of menthol with lauric acid (C12) was found to conform to a ping-pong bi-bi mechanism with dead-end inhibition by excess menthol. According to the mechanism we determined the maximum reaction velocity (V(max)), Michaelis constants (K(m(fatty acid) and K(m(menthol)), and the inhibition constant (K(i)). Results have shown that the constant values K(m) and K(i) of (+)-menthol are comparable with the corresponding values of the (-)-isomer. However, there was significant difference in the V(max) values between the two enantiomers; the V(max) involving the (-)-isomer was a 100-fold faster than that of the (+)-isomer. The results suggest that the lipase should recognize the chirality of menthol molecule not in the binding process to the hydrophobic pocket of the lipase but in the nucleophilic attack of the OH group in (-)-menthol. To assess the substrate specificity of C. rugosa lipase for carboxylic acids, the effect of fatty acids of varying chain lengths (C14, C16) on esterification kinetics have been studied using the (-)-menthol. We found that fatty acids exhibit different K(m) values, whereas their V(max) values are invariably similar. The results have indicated that the catalytic activity of the surfactant-coated lipase for different fatty acid largely depends on the binding behavior of a fatty acid to the active site of the coated enzyme and this action is independent of the acylation step of the (-)-isomer by an acyl-enzyme intermediate.. |
| 555. |
Shin Ya Okazaki, Noriho Kamiya, Kojiro Abe, Masahiro Goto, Fumiyuki Nakashio, Novel preparation method for surfactant-lipase complexes utilizing water in oil emulsions, Biotechnology and Bioengineering, 10.1002/(SICI)1097-0290(19970720)55:23.0.CO;2-E, 55, 2, 455-460, 1997.07, A novel preparation method for surfactant-lipase complexes has been developed utilizing water in oil emulsions. In order to optimize the preparation conditions, we have investigated the effects of several operational parameters on the enzymatic activity of the surfactant-lipase complexes in organic media. When a nonionic surfactant was employed under optimal preparation conditions [alkaline pH 8-10, organic/aqueous = 90/10 (v/v), concentration of surfactant, 10 mM], the surfactant-lipase complex efficiently catalyzed the esterification of benzyl alcohol with lauric acid in organic media. The esterification rate of the surfactant-lipase complex was increased over 16-fold relative to the native powder lipase. Furthermore, the lipase complex showed high storage stability.. |
| 556. |
N Kamiya, M Goto, How is enzymatic selectivity of menthol esterification catalyzed by surfactant-coated lipase determined in organic media?, BIOTECHNOLOGY PROGRESS, 10.1021/bp9700317, 13, 4, 488-492, 1997.07, A kinetic study of menthol esterification reaction catalyzed by surfactant-coated lipase (from Candida rugosa) has been conducted in dry isooctane. The kinetic characteristics observed in the enantioselective esterification of menthol with lauric acid (C-12) was found to conform to a ping-pong bi-bi mechanism with dead-end inhibition by excess menthol. According to the mechanism we determined the maximum reaction velocity (V-max), Michaelis constants (K-m(fatty acid) and K-m(menthol)), and the inhibition constant (K-i). Results have shown that the constant values K-m and K-i of (+)-menthol are comparable with the corresponding values of the (-)-isomer. However, there was significant difference in the V-max values between the two enantiomers; the V-max involving the (-)-isomer was a 100-fold faster than that of the (+)-isomer. The results suggest that the lipase should recognize the chirality of menthol molecule not in the binding process to the hydrophobic pocket of the lipase but in the nucleophilic attack of the OH group in (-)-menthol. To assess the substrate specificity of C. rugosa lipase for carboxylic acids, the effect of fatty acids of varying chain lengths (C-14, C-16) on esterification kinetics have been studied using the (-)-menthol. We found that fatty acids exhibit different K-m values, whereas their V-max values are invariably similar. The results have indicated that the catalytic activity of the surfactant-coated lipase for different fatty acid largely depends on the binding behavior of a fatty acid to the active site of the coated enzyme and this action is independent of the acylation step of the (-)-isomer by an acylenzyme intermediate.. |
| 557. |
SY Okazaki, N Kamiya, K Abe, M Goto, F Nakashio, Novel preparation method for surfactant-lipase complexes utilizing water in oil emulsions, BIOTECHNOLOGY AND BIOENGINEERING, 10.1002/(SICI)1097-0290(19970720)55:23.0.CO;2-E, 55, 2, 455-460, 1997.07, A novel preparation method for surfactant-lipase complexes has been developed utilizing water in oil emulsions. In order to optimize the preparation conditions, we have investigated the effects of several operational parameters on the enzymatic activity of the surfactant-lipase complexes in organic media. When a nonionic surfactant was employed under optimal preparation conditions [alkaline pH 8-10, organic/aqueous = 90/10 (v/v), concentration of surfactant, 10 mM], the surfactant-lipase complex efficiently catalyzed the esterification of benzyl alcohol with lauric acid in organic media. The esterification rate of the surfactant-lipase complex was increased over 16-fold relative to the native powder lipase. Furthermore, the lipase complex showed high storage stability. (C) 1997 John Wiley & Sons, Inc.. |
| 558. |
S Noda, N Kamiya, M Goto, F Nakashio, Enzymatic polymerization catalyzed by surfactant-coated lipases in organic media, BIOTECHNOLOGY LETTERS, 10.1023/A:1018334430266, 19, 4, 307-309, 1997.04, Structural ring-opening of lactones driven by enzymatic polymerization has been performed using low concentration dosages of surfactant-coated lipases in organic media. By comparison, enzymatic polymerization rate with coated lipase proceeded at a rate 100-fold better than native powder. Similarly a higher polymeric molecular weight (21,300), narrow dispersity (Mw/Mn = 1.9) and better conversion (100%) were obtained following polyesterification tests with surfactant-coated lipase.. |
| 559. |
Sadafumi Noda, Noriho Kamiya, Masahiro Goto, Fumiyuki Nakashio, Enzymatic polymerization catalyzed by surfactant-coated lipases in organic media, Biotechnology letters, 10.1023/A:1018334430266, 19, 4, 307-310, 1997.01, Structural ring-opening of lactones driven by enzymatic polymerization has been performed using low concentration dosages of surfactant-coated lipases in organic media. By comparison, enzymatic polymerization rate with coated lipase proceeded at a rate 100-fold better than native powder. Similarly a higher polymeric molecular weight (21,300), narrow dispersity (Mw/Mn = 1.9) and better conversion (100%) were obtained following polyesterification tests with surfactant-coated lipase.. |
| 560. |
Noriho Kamiya, Shin Ya Okazaki, Masahiro Goto, Surfactant-horseradish peroxidase complex catalytically active in anhydrous benzene, Biotechnology Techniques, 10.1023/A:1018400302660, 11, 6, 375-378, 1997.01, A surfactant-horseradish peroxidase complex was prepared with a nonionic surfactant by utilizing water-in-oil emulsions. The complex was readily soluble in anhydrous benzene in which it proved to be an excellent biocatalyst for the oxidation of o-phenylenediamine using H2O2 or tert-butyl hydroperoxide as oxidants.. |
| 561. |
Masahiro Goto, Sadafumi Noda, Noriho Kamiya, Fumiyuki Nakashio, Enzymatic resolution of racemic ibuprofen by surfactant-coated lipases in organic media, Biotechnology letters, 10.1007/BF00127899, 18, 7, 839-844, 1996.07, Surfactant-coated lipases have been utilized as a biocatalyst for the resolution of racemic ibuprofen. S-(+)-ibuprofen was selectively transferred to the ester form by Mucor javanicus or Candida rugosa lipase. The enzymatic activity of lipases in organic media was remarkably enhanced by coating with a nonionic surfactant. The reaction rates of the coated lipases were increased around 100-fold that of the powder lipases.. |
| 562. |
M Goto, S Noda, N Kamiya, F Nakashio, Enzymatic resolution of racemic ibuprofen by surfactant-coated lipases in organic media, BIOTECHNOLOGY LETTERS, 10.1007/BF00127899, 18, 7, 839-844, 1996.07, Surfactant-coated lipases have been utilized as a biocatalyst for the resolution of racemic ibuprofen. S-(+)-ibuprofen was selectively transferred to the ester form by Mucor javanicus or Candida rugosa lipase. The enzymatic activity of lipases in organic media was remarkably enhanced by coating with a nonionic surfactant. The reaction rates of the coated lipases were increased around 100-fold that of the powder lipases.. |
| 563. |
Noriho Kamiya, Emiko Murakami, Masahiro Goto, Fumiyuki Nakashio, Effect of using a co-solvent in the preparation of surfactant-coated lipases on catalytic activity in organic media, Journal of Fermentation and Bioengineering, 10.1016/0922-338X(96)89451-7, 82, 1, 37-41, 1996.01, Surfactant-coated lipases (from Candida cylindracea) were prepared in phosphate buffer (pH 6.9) containing a 2% (v/v) water-miscible organic solvent. The molecular structure of the surfactants used to coat the lipase strongly affected the esterification activity of the modified lipases in isooctane, while both the yield and the enzymatic activity were significantly influenced by the nature of the co-solvent used during the preparation. Although, in general, the most suitable co-solvents were found to be hydrophilic, especially ethanol and N,N-dimethylformamide (DMF), no definite correlation between solvent hydrophobicity (log P) and the properties of the surfactant-coated lipases was revealed. However, when a co-solvent which can dissolve a surfactant well was employed, the surfactant number attached to one lipase molecule tended to decrease and the yield was also lowered. Addition of dimethylsulfoxide (DMSO), formamide, or ethylene glycol to the lipase buffer solution during preparation led to substantial decreases in catalytic activity, even though the modified lipases were obtained in a good yield. This results suggests that these co-solvents affect the three- dimensional structure of the lipase, that is, the catalytic activity of surfactant-coated lipases in organic media may be chiefly determined during the preparation process in an aqueous solution.. |
| 564. |
Masahiro Goto, Masaki Miyata, Noriho Kamiya, Fumiyuki Nakashio, Novel surfactant-coated enzymes immobilized in poly(ethylene glycol) microcapsules, Biotechnology Techniques, 10.1007/BF00224402, 9, 2, 81-84, 1995.02, Novel surfactant-coated enzymes immobilized in poly(ethylene glycol) microcapsules have been developed for the re-use of an oil-soluble enzyme in organic media. The esterification rate of the surfactant-coated lipase immobilized in the microcapsules was thirty times that of the powder lipase. More than 90% of the enzymatic activity of the capsulated lipases has been maintained after recycling six times.. |
| 565. |
M GOTO, M MIYATA, N KAMIYA, F NAKASHIO, NOVEL SURFACTANT-COATED ENZYMES IMMOBILIZED IN POLY(ETHYLENE GLYCOL) MICROCAPSULES, BIOTECHNOLOGY TECHNIQUES, 10.1007/BF00224402, 9, 2, 81-84, 1995.02, Novel surfactant-coated enzymes immobilized in poly(ethylene glycol) microcapsules have been developed for the re-use of an oil-soluble enzyme in organic media. The esterification rate of the surfactant-coated lipase immobilized in the microcapsules was thirty times that of the powder lipase. More than 90 % of the enzymatic activity of the capsulated lipases has been maintained after recycling six times.. |
| 566. |
Masahiro Goto, Muneharu Goto, Noriho Kamiya, Fumiyuki Nakashio, Enzymatic interesterification of triglyceride with surfactant‐coated lipase in organic media, Biotechnology and Bioengineering, 10.1002/bit.260450105, 45, 1, 27-32, 1995.01, Several surfactant‐coated enzymes have been prepared by coating lipases of various origins with a nonionic surfactant, glutamic acid dioleylester ribitol (2C18Δ9GE). Enzymatic interesterification of tripalmitin with oleic acid using the surfactant‐coated lipase was carried out in organic media. The surfactant‐coated lipases could effectively catalyze the interesterification of glycerides better than did the powder lipases. A suitable organic solvent was an aliphatic hydrocarbon such as isooctane. The enzymatic activity for the interesterification strongly depended on the origin of the lipase. The surfactant‐coated lipase prepared by Mucor javanicus showed the highest enzymatic activity for the interesterification of glycerides, although its powder lipase did not show enzymatic activity. Selective interesterification of glycerides could be performed by adjusting the concentration ratio of oleic acid to tripalmitin in isooctane. Di‐substituted glyceride could be selectively produced when the concentration ratio of carboxylic acid to glycerides was 7. © 1995 John Wiley & Sons, Inc.. |
| 567. |
Noriho Kamiya, Masahiro Goto, Fumiyuki Nakashio, Surfactant‐Coated Lipase Suitable for the Enzymatic Resolution of Menthol as a Biocatalyst in Organic Media, Biotechnology Progress, 10.1021/bp00033a005, 11, 3, 270-275, 1995.01, Enantioselective esterification of menthol with fatty acids using a surfactant‐coated lipase was carried out in organic media. The surfactant‐coated lipase originating from Candida cylindracea appeared to be highly enantioselective and good biocatalyst for the resolution of racemic menthol. The enzymatic activity of the lipase in organic media was significantly increased by a coating with a nonionic surfactant. The reaction rate of the coated lipase was more than 100 times that of the powder lipase. In order to investigate the effect of the organic solvent on enantioselectivity, 19 kinds of solvents were employed. The nature of the organic solvent strongly affected the efficiency of the biocatalyst and the enantioselectivity. Among them, isooctane was the best organic solvent from the viewpoint of reaction rate and enantioselectivity. The effect of reaction temperature on esterification was also investigated. The optimal reaction temperature was around 35 °C. The enzymatic activities using n‐saturated fatty acids with different alkyl chain lengths were compared, and long‐chain fatty acids were found to be better substrates than shorter ones. The relationship between the initial rate of the esterification and the carbon number of the fatty acid was not linear. These results suggest that there are inherent Km values for each fatty acid.. |
| 568. |
Masahiro Goto, Muneharu Goto, Muneharu Goto, Noriho Kamiya, Fumiyuki Nakashio, Enzymatic interesterification of triglyceride with surfactant‐coated lipase in organic media, Biotechnology and Bioengineering, 10.1002/bit.260450105, 45, 1, 27-32, 1995.01, Several surfactant‐coated enzymes have been prepared by coating lipases of various origins with a nonionic surfactant, glutamic acid dioleylester ribitol (2C18Δ9GE). Enzymatic interesterification of tripalmitin with oleic acid using the surfactant‐coated lipase was carried out in organic media. The surfactant‐coated lipases could effectively catalyze the interesterification of glycerides better than did the powder lipases. A suitable organic solvent was an aliphatic hydrocarbon such as isooctane. The enzymatic activity for the interesterification strongly depended on the origin of the lipase. The surfactant‐coated lipase prepared by Mucor javanicus showed the highest enzymatic activity for the interesterification of glycerides, although its powder lipase did not show enzymatic activity. Selective interesterification of glycerides could be performed by adjusting the concentration ratio of oleic acid to tripalmitin in isooctane. Di‐substituted glyceride could be selectively produced when the concentration ratio of carboxylic acid to glycerides was 7. © 1995 John Wiley & Sons, Inc. Copyright © 1995 John Wiley & Sons, Inc.. |
| 569. |
Masahiro Goto, Noriho Kamiya, Masaki Miyata, Fumiyuki Nakashio, Enzymatic Esterification by Surfactant‐Coated Lipase in Organic Media, Biotechnology Progress, 10.1021/bp00027a005, 10, 3, 263-268, 1994.01, Surfactant‐coated lipases have been prepared with a synthesized surfactant. Preparation conditions to obtain a suitable surfactant‐coated lipase were investigated. The enzymatic activity of the lipase in an organic solvent significantly increased with the coating of the surfactant. The esterification rate from the surfactant‐coated lipase was much higher than that from the powder lipase. An aliphatic solvent showed higher activity than did alcohol, aromatic, and chloric solvents. Among them, isooctane gave the highest activity. The reactivity of the surfactant‐coated lipase depends on the pH of the aqueous solution in the preparation and on the buffer solution. Surfactant‐coated lipase prepared in the middle pH range using phosphate buffer showed high enzymatic activity. The surfactant‐coated lipase was thermostable at high temperature compared to the native lipase. A kinetic study enabled a ping‐pong bi‐bi reaction mechanism with alcohol inhibition to be suggested. From the kinetic analysis, it was found that an alcohol substrate inhibits enzymatic esterification by lipase. The reaction rate of the coated lipase was about 100 times that of the powder lipase.. |
