Language：English   Publishing type：Research paper (scientific journal)   Publisher：NATURE PORTFOLIO  

The development of simple and safe methods for recovering environmental pollutants, such as heavy metals, is needed for sustainable environmental management. Short elastin-like peptide (ELP) analogues conjugated with metal chelating agents are considered to be useful as metal sequestering agents as they are readily produced, environment friendly, and the metal binding domain can be selected based on any target metal of interest. Due to the temperature dependent self-assembly of ELP, the peptide-based sequestering agents can be transformed from the solution state into the particles that chelate metal ions, which can then be collected as precipitates. In this study, we developed a peptide-based sequestering agent, AADAAC-(FPGVG)(4), by introducing the metal-binding sequence AADAAC on the N-terminus of a short ELP, (FPGVG)(4). In turbidity measurements, AADAAC-(FPGVG)(4) revealed strong self-assembling ability in the presence of metal ions such as Cd2+ and Zn2+. The results from colorimetric analysis indicated that AADAAC-(FPGVG)(4) could capture Cd2+ and Zn2+. Furthermore, AADAAC-(FPGVG)(4) that bound to metal ions could be readily recycled by treatment with acidic solution without compromising its metal binding affinity. The present study indicates that the fusion of the metal-binding sequence and ELP is a useful and powerful strategy to develop cost-effective heavy metal scavenging agents with low environmental impacts.

DOI： 10.1038/s41598-022-05695-w

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DOI： https://doi.org/ 10.1371/journal.pone.0246583

Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1021/acsomega.0c06140.

Language：English   Publishing type：Research paper (scientific journal)  

Halogenation of organic compounds is one the most important transformations in chemical synthesis and is used for the production of various industrial products. A variety of halogenated bisphenol analogs have recently been developed and are used as alternatives to bisphenol A (BPA), which is a raw material of polycarbonate that has adverse effects in animals. However, limited information is available on the potential toxicity of the halogenated BPA analogs. In the present study, to assess the latent toxicity of halogenated BPA analogs, we evaluated the binding and transcriptional activities of halogenated BPA analogs to the estrogen-related receptor γ(ERRγ), a nuclear receptor that contributes to the growth of nerves and sexual glands. Fluorinated BPA analogs demonstrated strong ERRγbinding potency, and inverse antagonistic activity, similar to BPA. X-ray crystallography and fragment molecular orbital (FMO) calculation revealed that a fluorine-substituted BPA analog could interact with several amino acid residues of ERRγ-LBD, strengthening the binding affinity of the analogs. The ERRγbinding affinity and transcriptional activity of the halogenated BPAs decreased with the increase in the size and number of halogen atom(s). The IC₅₀ values, determined by the competitive binding assay, correlated well with the binding energy obtained from the docking calculation, suggesting that the docking calculation could correctly estimate the ERRγbinding potency of the BPA analogs. These results confirmed that ERRγhas a ligand binding pocket that fits very well to BPA. Furthermore, this study showed that the binding affinity of the BPA analogs can be predicted by the docking calculation, indicating the importance of the calculation method in the risk assessment of halogenated compounds.

DOI： 10.1021/acs.chemrestox.9b00379

Language：English   Publishing type：Research paper (scientific journal)  

17β-Estradiol (E2) is a natural steroid ligand for the structurally and physiologically independent estrogen receptors (ERs) ERα and ERβ. We recently observed that CF₃-containing bisphenol AF (BPAF) works as an agonist for ERα but as an antagonist for ERβ. Similar results were also observed for the CCl₃-containing bisphenol designated as HPTE. Both BPAF and HPTE are comprised of a tri-halogenated methyl group in the central alkyl moiety of their bisphenol structures, which strongly suggests that halogens contribute directly to the agonist/antagonist dual biological functions. We conducted this study to investigate the structure-activity relationships by assessing together newly synthesized CF₃- and CBr₃-containing bisphenol E analogs (BPE-X). We first tested bisphenols for their receptor binding ability and then for their transcriptional activities. Halogen-containing bisphenols were found to be fully active for ERα, but almost completely inactive for ERβ. When we examined these bisphenols for their inhibitory activities for E2 in ERβ, we observed that they worked as distinct antagonists. The ascending order of agonist/antagonist dual biological functions was BPE-F < BPE-Cl (HPTE) ≤ BPAF < BPE-Br, demonstrating that the electrostatic halogen bonding effect is a major driving force of the bifunctional ERα agonist and ERβ antagonist activities of BPAF.

DOI： 10.1016/j.bmc.2019.115274

Language：English   Publishing type：Research paper (scientific journal)  

An endocrine-disrupting chemical Bisphenol A (BPA) binds specifically to a nuclear receptor (NR) named ERRγ. Although the importance of receptor-binding evaluation for human NRs is often stressed, the binding characteristics of so-called next-generation (NextGen) bisphenol compounds are still poorly understood. The ultimate objective of this investigation was to evaluate BPA and its NextGen analogs for their abilities to bind to 21 human NRs, the greatest members of NRs for which tritium-labeled specific ligands were available. After establishing the detailed assay conditions for each NR, the receptor binding affinities of total 11 bisphenols were evaluated in competitive binding assays. The results clearly revealed that BPA and the NextGen bisphenols of BPAF, BPAP, BPB, BPC, BPE, and BPZ were highly potent against one or more of NRs such as CAR, ERα, ERβ, ERRγ, and GR, with IC₅₀ values of 3.3–73 nM. These bisphenols were suggested strongly to be disruptive to these NRs. BPM and BPP also appeared to be disruptive, but less potently. BPF exhibited only weak effects and only against estrogen-related NRs. Surprisingly, most doubtful bisphenol BPS was supposed not to be disruptive. The NRs to which BPA and NextGen bisphenols did not bind were RARα, RARβ, RARγ, and VDR. PPARγ, RORα, RORβ, RORγ, RXRα, RXRβ, and RXRγ, exhibited very weak interaction with these bisphenols. The ten remaining NRs, namely, ERRγ, ERβ, ERα, CAR, GR, PXR, PR, AR, LXRβ, and LXRα, showed distinctly strong binding to some bisphenols in this order, being likely to have consequential endocrine-disruption effects.

DOI： 10.1016/j.taap.2019.114610

Language：English   Publishing type：Research paper (scientific journal)  

An endocrine-disrupting chemical Bisphenol A (BPA) binds specifically to a nuclear receptor (NR) named ERRγ. Although the importance of receptor-binding evaluation for human NRs is often stressed, the binding characteristics of so-called next-generation (NextGen) bisphenol compounds are still poorly understood. The ultimate objective of this investigation was to evaluate BPA and its NextGen analogs for their abilities to bind to 21 human NRs, the greatest members of NRs for which tritium-labeled specific ligands were available. After establishing the detailed assay conditions for each NR, the receptor binding affinities of total 11 bisphenols were evaluated in competitive binding assays. The results clearly revealed that BPA and the NextGen bisphenols of BPAF, BPAP, BPB, BPC, BPE, and BPZ were highly potent against one or more of NRs such as CAR, ERα, ERβ, ERRγ, and GR, with IC50 values of 3.3–73 nM. These bisphenols were suggested strongly to be disruptive to these NRs. BPM and BPP also appeared to be disruptive, but less potently. BPF exhibited only weak effects and only against estrogen-related NRs. Surprisingly, most doubtful bisphenol BPS was supposed not to be disruptive. The NRs to which BPA and NextGen bisphenols did not bind were RARα, RARβ, RARγ, and VDR. PPARγ, RORα, RORβ, RORγ, RXRα, RXRβ, and RXRγ, exhibited very weak interaction with these bisphenols. The ten remaining NRs, namely, ERRγ, ERβ, ERα, CAR, GR, PXR, PR, AR, LXRβ, and LXRα, showed distinctly strong binding to some bisphenols in this order, being likely to have consequential endocrine-disruption effects.

DOI： https://doi.org/10.1016/j.taap.2019.114610

Language：English   Publishing type：Research paper (scientific journal)  

Elastin-like peptides (ELP) consist of distinctive repetitive sequences, such as (VPGVG)_n, exhibit temperature-dependent reversible self-assembly (coacervation), and have been considered to be useful for the development of thermo-responsive materials. Further fundamental studies evaluating coacervative properties of novel nonlinear ELPs could present design concepts for new thermo-responsive materials. In this study, we prepared novel ELPs, cyclic (FPGVG)_n (cyclo[FPGVG]_n, n = 1-5), and analyzed its self-assembly properties and structural characteristics. Cyclo[FPGVG]_n (n = 3-5) demonstrated stronger coacervation capacity than the corresponding linear peptides. The coacervate of cyclo[FPGVG]₅ was able to retain water-soluble dye molecules at 40°C, which implied that cyclo[FPGVG]₅ could be employed as a base material of DDS (Drug Delivery System) matrices and other biomaterials. The results of molecular dynamics simulations and circular dichroism measurements suggested that a certain chain length was required for cyclo[FPGVG]_n to demonstrate alterations in molecular structure that were critical to the exhibition of coacervation.

DOI： 10.1021/acs.biomac.8b00353

Language：English   Publishing type：Research paper (scientific journal)  

The elastin-derived peptide analog, (FPGVG)5, demonstrates a temperature- dependent self-association (coacervation) property. In this study, we prepared cyclic (FPGVG)n (n=1-5) analogs and investigated their coacervation and structural properties to assess their potential for use as a biomaterial. We found that the cyclic (FPGVG)n (n=3-5) analogs exhibited high coacervation ability. The results from the molecular dynamics simulation suggest that turn structures were important for coacervation of elastin-derived cyclic peptide analogs.

Language：English   Publishing type：Research paper (other academic)  

The elastin-derived peptide analog, (FPGVG)5, demonstrates a temperature- dependent self-association (coacervation) property. In this study, we prepared fluorescent-labeled (FPGVG)5 analogs, and evaluated their coacervation and fluorescent properties in order to assess their potential for use as a base material for fluorescent thermometers. We found that the fluorescent-labeled (FPGVG)5 analogs exhibited high coacervation ability. Results from the molecular dynamics calculation suggest that the fluorescent group is exposed on the surface of the molecule.

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A series of elastin-derived peptide (Phe-Pro-Gly-Val-Gly)5 dimers possessing high coacervation potential were synthesized. The new dimeric peptides showed significantly high coacervation ability compared to known elastin-derived peptide analogs. The molecular dynamics calculation results reveal that the dimeric peptides contain characteristic sheet-turnsheet motif involving a type II β-turn-like structure and form globular conformation.

DOI： 10.1002/psc.2876

Language：English   Publishing type：Research paper (other academic)  

A tryptophan-containing elastin-derived dimeric peptide (C(WPGVG)3)2 shows potent self-assembly activity. To elucidate the underlying coacervation mechanisms, we measured the fluorescence of (C(WPGVG)3)2 by using 1,8-ANS as a fluorescent probe. The results revealed that the peptide forms microaggregates at temperature and concentration lower than those at which visible coacervation occurs. These findings suggest that microaggregate formation is unrelated to the maturation of the coacervate drop, which is temperature-dependent.

Language：Japanese   Publishing type：Research paper (international conference proceedings)  

It has been shown that elastin-derived peptides containing aromatic amino acids, (FPGVG)n and (WPGVG)n, demonstrate coacervation at small repetition numbers (n = 3–5). In this study, we prepared dimer peptides of (FPGVG)5 and (WPGVG)3 and evaluated their coacervation properties. The coacervation ability of the peptides was significantly enhanced by the dimerization. The results of molecular dynamics calculation suggest that the sheet-turn-sheet motif is important for the self-assembly of FPGVG-related peptides.

Language：Japanese   Publishing type：Research paper (international conference proceedings)  

Elastin is the core protein of elastic fibers in the elastic tissues, such as arterial walls, lungs, and skin, and plays an essential role in tissue biomechanics, providing the extensibility and condensability. Elastin shows self-association, which is referred as coacervation, under physiological conditions. Coacervation property of elastin characterized by reversible association/dissociation phenomena is considered to be important for its elasticity. Tropoelastin, the precursor protein of elastin, contains characteristic repetitive sequences in its hydrophobic regions. The pentapeptide sequence, Val-Pro-Gly-Val-Gly (VPGVG), is one of the repeating sequences, and presents in almost all mammal species analyzed so far. To clarify the mechanism of elastin’s elasticity, many studies about coacervation properties of elastin-derived peptide-analogs have been conducted. Many previous studies reported that large molecular size was needed for coacervation of elastin-derived peptides. For instance, (VPGVG)n demonstrated the coacervation property only when they had large numbers of repetition (n>40). Recently, we developed the synthetic hydrophobic oligomers (IPGVG)n and (FPGVG)n, and reported that they demonstrate coacervation at significantly smaller repetition numbers (n=5~7). Because of their low molecular size, these analogs are expected to be useful as model peptides for structure analysis and base materials for developing various biomedical products, skin substitutes, synthetic vascular grafts, and drug delivery systems.
In this study, to obtain more short coacervatable peptides, we synthesized dimers of pentapeptide (FPGVG)5 and investigated their coacervation property and secondary structures. To produce dimer peptides, cysteine-conjugated pentapeptide Cys-(FPGVG)5 and (FPGVG)5-Cys were synthesized by the solid-phase method using Fmoc strategy. By aerobic oxidation reaction of each Cys-peptide, homodimers, linked by disulfide bond, were obtained. Synthesized peptides were purified by HPLC and analyzed by MALDI-TOF- MS. The dimer peptides clearly showed coacervation around 15˚C in the condition at 10 mg/ml, whereas (FPGVG)5 required rather higher temperature and at least 20 mg/ml of peptide to demonstrate coacervation. These results indicated that the dimerization of elastin-derived peptide-analogs significantly enhanced the coacervation ability of (FPGVG)5. Furthermore, these dimer peptides showed reversible temperature-dependent conformation change in CD measurement.

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DOI： 10.1002/anie.200905158

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DOI： 10.3987/COM-08-S(F)93

Language：English   Publishing type：Research paper (scientific journal)   Publisher：Biochemistry  

Synthetic elastin-like peptides (ELPs) that possess characteristic tropoelastin-derived hydrophobic repetitive sequences, such as (VPGVG)n, exhibit thermoresponsive reversible self-assembly. Although their thermoresponsive properties have been well-studied, the sequence-dependent and structural requirements for self-assembly remain ambiguous. In particular, it is still unclear whether the amino acid sequences derived from tropoelastin are necessary for self-assembly. In this study, 11 sequence-shuffled ELP analogues based on (FPGVG)5, which is a previously developed short ELP (sELP), were designed to elucidate the sequence-dependent and structural requirements for their self-assembly. Among them, eight shuffled peptides exhibited self-assembling properties, whereas the other three peptides were difficult to dissolve in water. Structural analyses revealed that the structural characteristics of the three insoluble peptides were different from those of their thermoresponsive analogues. Furthermore, the secondary structures of the peptide analogues possessing the self-assembly abilities were different from each other. These results suggest that the potential for self-assembly and water solubility of sELPs depend on the primary structure in each repeated unit. Moreover, several shuffled analogues exhibited more potent self-assembling properties than the original (FPGVG)5, indicating that shorter ELPs can be obtained using their novel motifs as repetitive units. We also observed that the presence of Pro-Gly sequence in the repeating units was advantageous in terms of peptide solubility. Although further analysis will be necessary to elucidate the molecular mechanism underlying the self-assembly of these sELPs, this study provides insights into the relationship between the amino acid sequence and the self-assembling ability of the peptides for developing new sELPs for various applications.

DOI： 10.1021/acs.biochem.3c00146

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Language：Japanese   Publisher：Faculty of Arts and Science, Kyushu University  

"Experimental approach to Natural Science" is a course that aims to deepen students' interest and understanding of natural science through experiments. The class was developed with "liberal science" in mind and was first offered in the summer quarter of 2022. After all classes were completed, we conducted an interview survey of the students who took the class, asking whether the class met his/her expectations and whether the class changed his/her view of natural science. The results of the survey revealed: (1) Most of the students had expected to conduct many experiments and to learn actively in this class, and the class generally met their expectations; (2) Students became more familiar with natural science through the class and showed a positive attitude toward learning natural science. On the other hand, based on the students' attitude during the course and the results of the interviews, the following improvements could be made: (1) to emphasize training to logically construct and test hypotheses rather than acquiring new knowledge in this course; (2) to provide more detailed explanations regarding the purpose of each experiment and interpretation of the results.

DOI： 10.15017/6770262

CiNii Research

Language：English   Publishing type：Research paper (scientific journal)   Publisher：Heliyon  

Halogenated flame retardants comprising bisphenol A (BPA) derivatives, such as tetrabromobisphenol A (TBBPA), have been studied their adverse effects on human health. However, despite the fact that these halogenated BPAs are easily degraded in the environment, the risks to living organisms due to these degraded products have mostly been overlooked. To evaluate the potential toxicity of degraded TBBPAs and related compounds, we examined the cytotoxicity of halogenated bisphenol A derivatives possessing one to four halogen atoms in vitro. The results indicated that the degraded TBBPA derivatives exhibited strong cytotoxicity against HeLa cells than TBBPA. Interestingly, the di-halogenated BPA derivatives possessing two halogen atoms exhibited the strongest cytotoxicity among tested compounds. In addition, a lactate dehydrogenase release assay, fluorescence spectroscopy and flow cytometry results indicated that dibromo-BPA and diiodo-BPA induced both apoptotic and necrotic cell death by damaging the cell membranes of HeLa cells. Moreover, Escherichia coli growth was inhibited in the presence of dehalogenated TBBPA and related compounds. These findings suggest that halogenated BPA derivatives that leak from various flame-retardant-containing products require strict monitoring, as not only TBBPA but also its degraded products in environment can exert adverse effects to human health.

DOI： 10.1016/j.heliyon.2023.e13003

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：WILEY  

Elastin-like peptides (ELPs) exhibit a reversible phase transition, known as coacervation, triggered by temperature changes. This property makes them useful as stimuli-responsive molecular materials for various applications. Among ELPs, short peptide chain lengths have some advantages over long peptide chain lengths because short ELPs can be easily obtained by chemical synthesis, allowing the use of various amino acids, including D-type and unnatural amino acids, at any position in the sequence. Moreover, the incorporated amino acids readily affect the temperature-responsive behavior of ELPs. However, to be utilized in various applications, it is necessary to develop short ELPs and to investigate their temperature-responsive properties. To obtain further insights into the temperature-responsive behavior of the short ELPs, we investigated branched short ELP analogs composed of (FPGVG)(n) chains (n = 1 or 2, abbreviated as F1 and F2, respectively). We synthesized multimers composed of four F1 chains or two to four F2 chains using ethylenediaminetetraacetic acid (EDTA) as a central component of multimerization. Our results show that the multimers obtained exhibited coacervation in aqueous solutions whereas linear F1 or F2 did not. Furthermore, the structural features of the obtained multimers were the same as those of linear (FPGVG)(4). In this study, we demonstrated that molecules capable of coacervation can be obtained by multimerization of F1 or F2. The temperature-responsive molecules obtained using short ELPs make it possible to use them as easy-to-synthesize peptide tags to confer temperature responsiveness to various molecules, which will aid the development of temperature-responsive biomaterials with a wide variety of functions.

DOI： 10.1002/psc.3449

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CiNii Research

CiNii Research

CiNii Research

Language：English   Publishing type：Research paper (scientific journal)  

Bisphenol A and its derivatives are recognized as endocrine disruptors based on their complex effects on estrogen receptor (ER) signaling. While the effects of bisphenol derivatives on ER alpha have been thoroughly evaluated, how these chemicals affect ER beta signaling is less well understood. Herein, we sought to identify novel ER beta ligands using a radioligand competitive binding assay to screen a chemical library of bisphenol derivatives. Many of the compounds identified showed intriguing dual activities as both ER alpha agonists and ER beta antagonists. Docking simulations of these compounds and ER beta suggested that they bound not only to the canonical binding site of ER beta but also to the coactivator binding site located on the surface of the receptor, suggesting that they act as coactivator-binding inhibitors (CBIs). Receptor-ligand binding experiments using WT and mutated ER beta support the presence of a second ligand-interaction position at the coactivator-binding site in ER beta, and direct binding experiments of ER beta and a coactivator peptide confirmed that these compounds act as CBIs. Our study is the first to propose that bisphenol derivatives act as CBIs, presenting critical insight for the future development of ER signaling-based drugs and their potential to function as endocrine disruptors.

DOI： 10.1016/j.jbc.2021.101173

Language：English   Publishing type：Research paper (scientific journal)  

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DOI： https://doi.org/10.3390/s20174781

Language：English   Publishing type：Research paper (other academic)  

Elastin-like peptide (ELP) analogs demonstrate reversible temperature-dependent association (coacervation) under physiological conditions. Previously, we revealed that simply dimerized and cyclized short ELP analogs show stronger coacervation activity than monomeric and linear counterpart of them, respectively. In this study, we investigated concentration dependency of the transition temperature of nonlinear short ELPs to estimate their coacervation activity. It was revealed that the transition temperature of nonlinear ELPs could be described as the logarithmic function of peptide concentration. In addition, by using this quantitative relationship, it was able to compare and estimate the coacervation activity of artificial ELPs with different water solubility.

Language：English   Publishing type：Research paper (other academic)  

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Language：English   Publishing type：Research paper (scientific journal)  

Employment of appropriate receptor conformations as templates is essential for appropriate identification of the latent receptor binding ability of chemicals using in silico docking calculations. In this study, we performed docking calculations using a number of agonist- and antagonist-bound conformations of 83 estrogen receptor (ER) α-ligand-binding domains as templates to clarify the type of receptor conformations required for reasonable identification of endocrine disruptors. Our results showed that 17β-estradiol and diethylstilbestrol (ERα agonists) bound preferentially to the agonist conformations, whereas 4-hydroxytamoxifen and raloxifene (ERα antagonists) bound selectively to the antagonist conformations. We also observed that bisphenol A analogues, which are partial agonists, bound more moderately and preferentially to the agonist conformations as compared with the antagonist conformations. Additionally, the docking calculations were able to estimate biological agonist or antagonist activity of chemicals based on the receptor conformation selectivity. Furthermore, structural analyses of the ligand-binding domains and docking calculation utilizing C-terminal-truncated receptors indicated that the C-terminal regions of these domains were capable of discriminating agonists from nonagonists. These results suggest that both agonist- and antagonist-binding conformations of receptors are necessary to predict the binding affinity and biological activity of chemicals for docking calculation-based risk assessment. Furthermore, this in silico method can be beneficial for drug discovery because it is useful for rapid searching of ligands for receptors and preventing the side effects caused by unfavorable receptor binding.

DOI： 10.1021/acsomega.9b00050

Language：English   Publishing type：Research paper (other academic)  

The elastin-derived peptide analog, (FPGVG)n, demonstrates temperature-dependent self-association (coacervation) property. In this study, we revealed that simple dimerization and cyclization of (FPGVG)n analogs induced strong coacervation ability compared to the ability of monomeric linear (FPGVG)5.

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In search for enzyme inhibitors, we often encounter "promiscuous" enzyme inhibitors exhibiting nonspecific binding property toward enzyme active site. Therefore, inhibitory candidates should be mechanistically characterized as early as possible in discovery processes. However, there remains a lack of highly reliable and readily available methodology to evaluate specificity of initial hits inhibitors. The present study developed and established a novel DMSO-perturbing assay to identify promiscuous enzyme inhibitors. The assay successfully identified nonspecific binding inhibitors with a broad scope, typically by the attenuation of inhibitory activity by the influence of DMSO-addition. This attenuation would be attributed to the nonspecific binding property of inhibitors toward both productive and nonproductive (nondenatured) states of enzymes in perturbation solution. This working hypothesis was supported by spectroscopic analyses of enzyme conformations and analyses of solvent effects on perturbation. Overall, these results provided a novel concept of the DMSO-perturbing assay.

DOI： 10.1021/acsmedchemlett.9b00093

Language：English   Publishing type：Research paper (scientific journal)  

Elastin-like peptides (ELPs) are distinct, repetitive, hydrophobic sequences, such as (VPGVG)n, that exhibit coacervation, the property of reversible, temperature-dependent self-association and dissociation. ELPs can be found in elastin and have been developed as new scaffold biomaterials. However, the detailed relationship between their amino acid sequences and coacervation properties remains obscure because of the structural flexibility of ELPs. In this study, we synthesized a novel, dimeric ELP analogue (H-C(WPGVG)3-NH2)2, henceforth abbreviated (CW3)2, and analyzed its self-assembly properties and structural factors as indicators of coacervation. Turbidity measurements showed that (CW3)2 demonstrated coacervation at a concentration much lower than that of its monomeric form and another ELP. In addition, the coacervate held water-soluble dye molecules. Thus, potent and distinct coacervation was obtained with a remarkably short sequence of (CW3)2. Furthermore, fluorescence microscopy, dynamic light scattering, and optical microscopy revealed that the coacervation of (CW3)2 was a stepwise process. The structural factors of (CW3)2 were analyzed by molecular dynamics simulations and circular dichroism spectroscopy. These measurements indicated that helical structures primarily consisting of proline and glycine became more disordered at high temperatures with concurrent, significant exposure of their hydrophobic surfaces. This extreme change in the hydrophobic surface contributes to the potent coacervation observed for (CW3)2. These results provide important insights into more efficient applications of ELPs and their analogues, as well as the coacervation mechanisms of ELP and elastin.

DOI： 10.1021/acs.biochem.7b01144

Language：English   Publishing type：Research paper (scientific journal)  

Many types of insects have been used as foods and protein sources. In this study, we investigated the usefulness of housefly larvae (Musca domestica) based on their amino acid composition and multifunctional biological activities. First, the utility of the amino acid composition of housefly larvae was evaluated by amino acid analysis. Notably, the housefly larvae contained sufficient amounts of all essential amino acids, and the amino acid composition was similar to that of hen eggs. Second, we prepared housefly larvae water extract (HLWE) using the decoction method and explored the biological activities of the extract for potential application of the extract as a functional food. HLWE showed significant antioxidant activity (75.4% at 5.00 mg/mL), angiotensin-I-converting enzyme (ACE) inhibitory activity (half-maximal inhibitory concentration [IC
₅₀
] = 0.430 mg/mL), and dipeptidyl peptidase-IV (DPP-IV) inhibitory activity ([IC
₅₀
] = 3.52 mg/mL). We found that the low-molecular-weight constituents (<6 kDa) in HLWE contributed to antioxidant and ACE-inhibitory activities, whereas the high-molecular-weight constituents (>6 kDa) contributed to DPP-IV inhibition. Our results suggested that housefly larvae may provide a useful source of multifunctional protein.

DOI： 10.1016/j.phanu.2017.09.001

Event date： 2023.3

Language：English  

Venue：東京理科大学野田キャンパス   Country：Japan  

Event date： 2022.11

Language：Japanese  

Venue：名古屋国際会議場   Country：Japan  

Event date： 2022.10

Language：English  

Venue：トークネットホール仙台   Country：Japan  

Event date： 2022.6

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：オンライン発表   Country：Japan  

Event date： 2021.11

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：オンライン発表   Country：Japan  

Event date： 2021.10

Language：English   Presentation type：Oral presentation (general)  

Venue：オンライン発表   Country：Japan  

Event date： 2020.11

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：オンライン開催   Country：Japan  

Event date： 2020.3

Language：English   Presentation type：Oral presentation (general)  

Venue：東京理科大学　野田キャンパス   Country：Japan  

Elastin-like peptides (ELPs), which consist of distinct repetitive sequences of elastin, exhibit temperature-dependent reversible self-assembly (coacervation) under physiological conditions. Due to this characteristic property, ELPs have been considered useful for the development of thermoresponsive biomaterials. Among ELPs, the pentapeptide sequence Val-Pro-Gly-Val-Gly (VPGVG) is the representative motif that exhibits coacervation. By mimicking this repetitive pentapeptide sequence, we have developed an elastin-derived short synthetic peptide, (FPGVG)n, that demonstrates stronger coacervation than (VPGVG)n. Although this peptide is potentially useful for developing stimuli-responsive biomaterials, the effects of sequential modification of ELPs on their coacervation activity are still unclear. In this study, (FPGVG)n-based peptide analogs whose phenylalanine residues were substituted with other amino acids were synthesized, and their coacervation properties were analyzed to elucidate the effect of the sequence on the self-association property.
The elastin-derived peptide (FPGVG)5 (F5) and F5-derived peptide analogs were successfully synthesized by the Fmoc solid phase method. Subsequently, the temperature-dependent coacervation properties of the synthetic peptides were evaluated by turbidity measurement. As a result, F5-analogs that contain aromatic amino acids (Tyr or Trp) as a substituent exhibited strong coacervation activity. When one of the Phe residues of F5 were substituted by a non-aromatic amino acid, the coacervation activity of the peptide significantly decreased, even if the replacement was a hydrophobic aliphatic amino acid. It was suggested that the coacervation activity of short ELPs is not simply determined by the hydrophobicity of the peptide and that interaction via aromatic amino acid residues was important for the coacervation activity of the peptides. It was also revealed that the coacervation activity of F5 was drastically affected by the substitution of a single amino acid residue.

Event date： 2019.10

Language：English   Presentation type：Oral presentation (general)  

Venue：東京医科歯科大学   Country：Japan  

Elastin-like peptide (ELP) analogs demonstrate reversible temperature-dependent association (coacervation) under physiological conditions. Previously, we revealed that simply dimerized and cyclized short ELP analogs show stronger coacervation activity than monomeric and linear counterpart of them, respectively. In this study, we investigated concentration dependency of the transition temperature of nonlinear short ELPs to estimate their coacervation activity. It was revealed that the transition temperature of nonlinear ELPs could be described as the logarithmic function of peptide concentration. In addition, by using this quantitative relationship, it was able to compare and estimate the coacervation activity of artificial ELPs with different water solubility.

Event date： 2019.9

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：パシフィコ横浜   Country：Japan  

弾性タンパク質エラスチンは、温度を上げると会合し下げると再び解離するという可逆的な自己集合（コアセルベーション）を示す。エラスチンは疎水性ドメインにVal-Pro-Gly-Val-Gly (VPGVG)のような特徴的な繰り返し配列を持ち、この繰り返し配列がコアセルベーションに重要であることが報告されている。また、この繰り返し配列を模倣したエラスチン由来ペプチド（Elastin-like Peptides; ELP）も同様に自己集合能を示すことから、人工血管や薬物送達システム等の基材として応用が可能なバイオマテリアル用の素材としての応用が期待されている。しかし、これらのペプチドがコアセルベーションを示す際の詳細な構造や機能、および配列に含まれる各アミノ酸残基の役割には不明な点が多く、分子素材としてELPを合理的にデザインするために、自己集合機構の解明が求められている。
代表的なELPである(VPGVG)n繰り返し配列ペプチドが自己集合を示すためには、200アミノ酸残基以上（配列繰り返し回数n>40）からなるポリペプチドが必要である。当研究室の先行研究では、繰り返し配列の先頭のVal残基をより疎水性の高いフェニルアラニン（Phe）に置換した合成ペプチド(FPGVG)nが25残基程度（n=5）の小分子でコアセルベーションを示すことを報告した。このことから、ELPがコアセルベーションを示すためには、ペプチドを構成する残基の疎水性が大きく関与していると考えられた。ところが、これの検証研究において、(FPGVG)5の中の１つの繰り返し配列における1個のPheを側鎖にヒドロキシ基を有するチロシン（Tyr）に置換したところ、親水性官能基が導入されたにも関わらずコアセルベーション能が高くなることが判明した。加えて、Tyrの置換位置の違いによってもコアセルベーション能が変化した。一方、Tyrと同様に側鎖にヒドロキシ基を有するセリン（Ser）でPheを置換すると、置換する位置に関わらず自己集合能が著しく低下した。これらの結果から、ELPの自己集合能は単純にアミノ酸の疎水性および親水性官能基の有無によって規定されるわけではないことが示唆された。そこで、さらにこれらの実験結果について分子構造から検証するため、分子動力学計算を用いて(FPGVG)5ならびにその一残基置換ペプチドの構造解析を行い、置換する残基の種類や位置によってペプチドの構造や分子内相互作用がどのように変化するか？について検証を行なった。

Event date： 2018.12

Language：English   Presentation type：Oral presentation (general)  

Venue：ロームシアター京都   Country：Japan  

Event date： 2016.10

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：京都テルサ   Country：Japan  

Event date： 2016.9

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：仙台国際センター   Country：Japan  

プラスチックの原料として幅広く使用されているビスフェノールA（BPA）が、核内受容体のひとつであるエストロゲン関連受容体γ型（ERRγ）に強く結合することが判明し、その機能を撹乱することが懸念されている。このため、代替としてBPAをハロゲンなどで置換した「新世代ビスフェノール」とよばれる一連の化合物が使用されるようになったが、ハロゲン化BPAもERRγに強く結合するともいわれており、これらの物質がもつ潜在的な撹乱リスクの評価が求められている。そこで本研究では、ハロゲン化がBPAのERRγに対する受容体結合性にどのように影響するかをin vitro とin silicoの手法により調べることを目的として実験を行った。
　ハロゲン化BPAとERRγの相互作用を解析するため、まずハロゲン化BPAを化学合成し、核内受容体ERRγに対する競合結合試験を行って結合親和性を調べた。その結果、ハロゲン置換基が大きく、また置換数が多くなるほどBPAアナログの結合親和性は低くなった。続いて、BPAアナログ10種について、5種類のカラムを用いた逆相HPLCを実施し、その溶出時間からリガンドの結合における相互作用の寄与を調べた。その結果、BPAアナログとカラム担体の相互作用は、主に疎水性相互作用が重要であること、また、π電子を介した相互作用を検証したところ、BPAの芳香環の電子とカラムのアルキル基の水素によるCH-π相互作用が主であることが示唆された。これらの相互作用がBPAとERRγの相互作用においても重要であると考えられた。また、ERRγの結晶構造（PDB ID: 2E2R）を鋳型として各種ハロゲン化BPAアナログのドッキング計算を行い、結合エネルギーが最小となるリガンドの最安定構造を用いてハロゲン化BPAアナログとERRγの結合構造を解析した。その結果、ハロゲン化BPAはBPAとほぼ同じ配向でERRγに結合するが、ヨウ素一置換体のみ立体障害の影響から他のアナログと異なる結合性を示し、その結果、生理活性が変わる可能性があることが示唆された。

Event date： 2016.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：同志社大学京田辺キャンパス   Country：Japan  

Self-assembly properties and structural analyses of fluorescent-labeled analogs of elastin-derived synthetic peptide (FPGVG)5

Event date： 2015.12

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Coacervation and fluorescence properties of fluorescent-labeled elastin-derived peptides

Event date： 2015.11

Language：English   Presentation type：Oral presentation (general)  

Venue：平塚中央公民館   Country：Japan  

Biological reactions within living cells are affected by temperature. Therefore, mapping the intracellular temperature of living cells is expected to contribute to an improved understanding of cellular events and the establishment of novel therapeutics. Previously, Uchiyama et al. demonstrated intracellular temperature mapping based on a fluorescent polymeric thermometer consisting mainly of acrylamide derivatives. Elastin-derived peptides exhibit reversible self-association (coacervation) under physiological conditions. Therefore, it was considered that these peptides might serve as useful materials for the development of such a fluorescent thermometer. Recently, we reported that the short synthetic elastin-derived peptide (FPGVG)5 demonstrates coacervation, which suggests that fluorescent-labeled (FPGVG)5 peptides, in particular, may be useful as basic materials for developing a novel fluorescent peptide-based thermometer. In the present study, we synthesized fluorescent-labeled analogs of (FPGVG)5 and investigated their coacervation property and peptide conformation.

Event date： 2015.11

Language：Japanese   Presentation type：Public lecture, seminar, tutorial, course, or other speech  

Venue：九州大学馬出キャンパス   Country：Japan  

Event date： 2015.5

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：九州大学箱崎キャンパス   Country：Japan  

Event date： 2015.3

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2014.10

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2014.10

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2014.5

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2014.3

Language：English   Presentation type：Oral presentation (general)  

Venue：つくば国際会議場（茨城県）   Country：Japan  

Event date： 2014.3

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2013.11

Language：English   Presentation type：Oral presentation (general)  

Venue：ホテル阪急エキスポパーク（大阪府）   Country：Japan  

Elastin is the core protein of elastic fibers in the elastic tissues plays an essential role in tissue biomechanics, providing the extensibility and condensability. Elastin shows coacervation (reversible association/dissociation phenomena) under physiological conditions. Tropoelastin, the precursor protein of elastin, contains characteristic repetitive sequences such as the pentapeptide sequence, Val-Pro-Gly-Val-Gly (VPGVG), in its hydrophobic regions. However, it was reported that large molecular size was needed for coacervation of elastin-derived peptide (VPGVG)n (numbers of repetition: n>40). Recently, we reported that synthetic hydrophobic oligomers (IPGVG)n and (FPGVG)n demonstrate coacervation at significantly smaller repetition numbers (n=5~7). Because of their low molecular size, these analogs are expected to be useful as model peptides for structure analysis and base materials for developing various biomedical products. In this study, to obtain more short coacervatable peptides, we synthesized dimers of pentapeptide (FPGVG)5 and investigated their coacervation property and secondary structures.

Event date： 2013.6

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：東京医科歯科大学（東京都）   Country：Japan  

Event date： 2013.3

Language：Japanese  

Country：Japan  

Event date： 2012.12

Language：Japanese  

Country：Japan  

Event date： 2012.12

Language：Japanese  

Country：Japan  

Event date： 2012.12

Language：Japanese  

Country：Japan  

Event date： 2012.9

Language：Japanese  

Country：Japan  

Event date： 2012.5

Language：Japanese  

Country：Japan  

Event date： 2012.3

Language：Japanese  

Country：Japan  

Event date： 2011.9

Language：Japanese  

Country：Japan  

Event date： 2011.9

Language：Japanese  

Country：Japan  

Event date： 2010.12 - 2016.12

Language：Japanese  

Country：Japan  

Event date： 2010.12

Language：Japanese  

Country：Japan  

Event date： 2010.8 - 2010.9

Language：Japanese  

Country：Japan  

Event date： 2010.5

Language：Japanese  

Country：Japan  

Event date： 2009.5

Language：English  

Country：Korea, Republic of  

Event date： 2008.11

Language：Japanese  

Country：Japan  

Event date： 2008.3

Language：Japanese  

Country：Japan  

Event date： 2023.10 - 2023.11

Language：Japanese  

Country：Japan  

Event date： 2021.11

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2021.10

Language：English   Presentation type：Oral presentation (general)  

Venue：オンライン発表   Country：Japan  

Event date： 2021.6

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2021.6

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：オンライン発表   Country：Japan  

Event date： 2020.5

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：誌上開催   Country：Japan  

Event date： 2019.6

Language：Japanese  

Venue：長崎大学文教キャンパス   Country：Japan  

Event date： 2019.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：甲南大学　岡本キャンパス   Country：Japan  

Event date： 2018.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：日本大学理工学部船橋キャンパス   Country：Japan  

Event date： 2017.12

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：神戸大学六甲台第2キャンパス   Country：Japan  

Event date： 2017.5

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：宮日会館/宮崎大学清武キャンパス   Country：Japan  

Event date： 2017.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：慶應義塾大学日吉キャンパス   Country：Japan  

The elastin-derived synthetic peptide analogs (FPGVG)n and (WPGVG)n, containing aromatic amino acids, demonstrate temperature-dependent self-association (coacervation). In this study, we prepared cyclic analogs of these elastin-derived peptides to investigate the molecular mechanism of coacervation and to obtain potent coacervatable peptides. It was revealed that cyclized elastin-derived peptide analogs, namely cyclic (FPGVG)n with n = 3–5, exhibited coacervation at a significantly lower concentration (1.0 mg/ml) than did linear (FPGVG)5 (requiring 30 mg/ml to show coacervation). Our results from molecular dynamics simulations suggest that local turn structures are important for coacervation of elastin-derived cyclic peptide analogs. In addition, our results from microscopy and dynamic light scattering measurements of cyclic (FPGVG)5 aqueous solutions indicate that this peptide can form liposome-like aggregate structures approximately 3 µm in diameter.

Event date： 2016.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：同志社大学京田辺キャンパス   Country：Japan  

Event date： 2016.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：同志社大学京田辺キャンパス   Country：Japan  

Event date： 2016.3 - 2015.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：同志社大学京田辺キャンパス   Country：Japan  

Event date： 2016.3 - 2015.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：同志社大学京田辺キャンパス   Country：Japan  

Event date： 2015.12

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：自治医科大学地域医療情報研修センター   Country：Japan  

Event date： 2015.12

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：神戸ポートアイランド   Country：Japan  

Event date： 2015.11

Language：English   Presentation type：Oral presentation (general)  

Venue：平塚中央公民館   Country：Japan  

Event date： 2015.6

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：北九州国際会議場   Country：Japan  

Event date： 2015.6

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：北九州国際会議場   Country：Japan  

Language：Japanese  

Language：Japanese  

Language：Japanese  

Language：Japanese  

Language：Japanese  

Type：Competition, symposium, etc. 

Number of participants：32

Type：Peer review 

Number of peer-reviewed articles in foreign language journals：2

Type：Competition, symposium, etc. 

Number of participants：49

Type：Competition, symposium, etc. 

Type：Competition, symposium, etc. 

Type：Competition, symposium, etc. 

Type：Competition, symposium, etc. 

Number of participants：50

Type：Competition, symposium, etc. 

Number of participants：68

2013年前期の自然科学総合実験は、当初の担当であった山田秀人助教が離任のため急遽担当した。

理学研究院化学部門構造機能生化学研究室の学生指導を委託され、学部学生２名、修士学生１名の実験指導を実施した（2014年3月まで）。