Updated on 2025/03/22

Information

 

写真a

 
MASUDA AKITSU
 
Organization
Faculty of Agriculture Department of Bioresource Sciences Assistant Professor
School of Agriculture Department of Bioresource and Bioenvironment(Concurrent)
Graduate School of Bioresource and Bioenvironmental Sciences Department of Bioresource Sciences(Concurrent)
Title
Assistant Professor
Contact information
メールアドレス
External link

Degree

  • Ph.D

Research Interests・Research Keywords

  • Research theme: Studies on functional modification of recombinant proteins produced in silkworms

    Keyword: Virus-like particles,antigen display,protein conjugation,protein stability

    Research period: 2024.1

  • Research theme: Production and functional evaluation of vaccine antigens using silkworm-baculovirus expression system

    Keyword: silkworm,baculovirus,vaccine,immunogenicity

    Research period: 2024.1

Papers

  • Production of norovirus VLPs of the nine representative genotypes widely distributed in Japan using the silkworm-baculovirus expression vector system

    Tsurumi Y., Morimoto K., Masuda A., Lee J.M., Mon H., Kusakabe T.

    Journal of Virological Methods   331   115038   2025.1   ISSN:01660934 eISSN:1879-0984

     More details

    Language:English   Publisher:Journal of Virological Methods  

    Norovirus (NoV) is one of the major causes of acute viral gastroenteritis in humans. Genetic variation is abundant, and prevalent genotypes vary from year to year and region to region. Since NoVs are difficult to amplify in cultured cells, genome RNA-free virus-like particles (VLPs) that mimic the capsid structure of the virus are promising vaccine candidates for the prevention of NoVs infection, and the development of multivalent VLP vaccines is required to prevent NoV infection in a wide range of genotypes. In this study, we attempted to produce NoV VLPs of the top nine genotypes that have a history of epidemics in Japan using the silkworm-baculovirus expression vector system (silkworm-BEVS), which has a proven track record in the mass production of recombinant proteins. In silkworm pupae infected with recombinant baculoviruses constructed to express VP1s, the major protein that forms VLP, the NoV VP1 protein was expressed in large amounts. Most genotypes of VP1 accumulated in the cytoplasm as soluble proteins, but solubility was reduced for that of two genotypes. VP1s of five genotypes could be purified in large quantities (>0.9 mg per pupa) by a two-step purification process, and gel filtration chromatography analysis confirmed the formation of VLPs. This study demonstrates the utility of silkworm-BEVS in producing NoV VLPs of multiple genotypes and provides the basis for the development of a multivalent vaccine against genetically diverse NoV infections.

    DOI: 10.1016/j.jviromet.2024.115038

    Web of Science

    Scopus

    PubMed

  • Diabetic Mice Spleen Vulnerability Contributes to Decreased Persistence of Antibody Production after SARS-CoV-2 Vaccine

    Atef, Y; Ito, T; Masuda, A; Kato, Y; Nishimura, A; Kanda, Y; Kunisawa, J; Kusakabe, T; Nishida, M

    INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES   25 ( 19 )   2024.10   ISSN:1661-6596 eISSN:1422-0067

     More details

    Language:English   Publisher:International Journal of Molecular Sciences  

    During the COVID-19 pandemic, diabetic and obese patients experienced higher rates of hospital admissions, severe illness, and mortality. However, vaccinations failed to provide those vulnerable populations the same level of protection against COVID-19 severity as those without diabetic and obese phenotypes. Our study aimed to investigate how diabetes mellitus (DM) impacts the immune response following vaccination including the artificially designed trimeric SARS-CoV-2 spike (S)-protein. By using two diabetic mouse models, ob/ob mice (obese, hyperglycemic, and insulin-resistant) and STZ-treated mice (insulin-deficient and hyperglycemic), we observed a significant reduction in S-protein-specific IgG antibody titer post-vaccination in both diabetic models compared to wild-type (WT) mice. Both diabetic mouse models exhibited significant abnormalities in spleen tissue, including marked reductions in splenic weight and the size of the white pulp regions. Furthermore, the splenic T-cell and B-cell zones were notably diminished, suggesting an underlying immune dysfunction that could contribute to impaired antibody production. Notably, vaccination with the S-protein, when paired with an optimal adjuvant, did not exacerbate diabetic cardiomyopathy, blood glucose levels, or liver function, providing reassurance about the vaccine′s safety. These findings offer valuable insights into potential mechanisms responsible for the decreased persistence of antibody production in diabetic patients.

    DOI: 10.3390/ijms251910379

    Web of Science

    Scopus

    PubMed

  • Effective expression and characterization of the receptor binding domains in SARS-CoV-2 Spike proteins from original strain and variants of concern using<i> Bombyx</i><i> mori</i> nucleopolyhedrovirus in silkworm

    Tsukamoto, A; Man, LJ; Oyama, K; Masuda, A; Mon, H; Ueda, T; Kusakabe, T

    PROTEIN EXPRESSION AND PURIFICATION   218   106450   2024.6   ISSN:1046-5928 eISSN:1096-0279

     More details

    Language:English   Publisher:Protein Expression and Purification  

    A new coronavirus, known as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is responsible for the global pandemic of COVID-19 in 2020. Through structural analysis, it was found that several amino acid residues in the human angiotensin-converting enzyme-2 (hACE2) receptor directly interact with those in the receptor binding domain (RBD) of the spike glycoprotein (S-protein). Various cell lines, including HEK293, HeLa cells, and the baculovirus expression vector system (BEVS) with the insect cell line Sf9, have been utilized to produce the RBD. In this study, we investigated the use of Bombyx mori nucleopolyhedrovirus (BmNPV) and BEVS. For efficient production of a highly pure recombinant RBD protein, we designed it with two tags (His tag and STREP tag) at the C-terminus and a solubilizing tag (SUMO) at the N-terminus. After expressing the protein using BmNPV and silkworm and purifying it with a HisTrap excel column, the eluted protein was digested with SUMO protease and further purified using a Strep-Tactin Superflow column. As a result, we obtained the RBD as a monomer with a yield of 2.6 mg/10 mL serum (equivalent to 30 silkworms). The RBD showed an affinity for the hACE2 receptor. Additionally, the RBDs from the Alpha, Beta, Gamma, Delta, and Omicron variants were expressed and purified using the same protocol. It was found that the RBD from the Alpha, Beta, Gamma, and Delta variants could be obtained with yields of 1.4–2.6 mg/10 mL serum and had an affinity to the hACE2 receptor.

    DOI: 10.1016/j.pep.2024.106450

    Web of Science

    Scopus

    PubMed

  • Characterization of high affinity IgM and IgG monoclonal antibodies against norovirus variants GII.4 and GII.17

    2024.5

     More details

  • SARS-CoV-2 strain-specific anti-spike IgG ELISA utilizing spike protein produced by silkworms

    Goto T., Sasaki T., Chong Y., Taniguchi M., Lee J.M., Masuda A., Ebihara T., Shiraishi K., Tani N., Yonekawa A., Gondo K., Kuwano H., Shimono N., Ikematsu H., Akashi K., Kusakabe T.

    Human Antibodies   31 ( 3 )   27 - 33   2023.9   ISSN:10932607

     More details

    Language:English   Publisher:Human Antibodies  

    BACKGROUND: A cost-effective and eco-friendly method is needed for the assessment of humoral immunity against SARS-CoV-2 in large populations. OBJECTIVE: We investigated the performance of an ELISA that uses silkworm-produced proteins to quantify the strain-specific anti-Spike IgG (anti-S IgG) titer. METHODS: The OD values for the anti-His-tag antibody, a standard material of ELISA quantification, were measured. Correlations between the ELISA for each strain and the Abbott SARS-CoV-2 IgG II Quant assay for the wild type were evaluated with serum samples from nine participants with various infection and vaccination statuses. RESULTS: Linear dose-responses were confirmed by high coefficients of determination: 0.994, 0.994, and 0.996 for the wild-type, Delta, and Omicron (BA.1) strain assays, respectively. The coefficient of determination for the wild-type and Delta strain assays was high at 0.959 and 0.892, respectively, while the Omicron strain assay had a relatively low value of 0.563. Booster vaccinees showed similar or higher titers against all strains compared to infected persons without vaccination. The Omicron-infected persons without vaccination had lower antibody titers against wild type than did the vaccinated persons. CONCLUSIONS: This study provides data indicating that the ELISA with silkworm-produced proteins makes it possible to discriminate and quantify the strain-specific anti-S IgG antibody induced by vaccination or infection.

    DOI: 10.3233/HAB-230006

    Scopus

    PubMed

  • Comprehensive Transcriptome Analysis in the Testis of the Silkworm, Bombyx mori

    Kakino, K; Mon, H; Ebihara, T; Hino, M; Masuda, A; Lee, JM; Kusakabe, T

    INSECTS   14 ( 8 )   2023.8   ISSN:2075-4450 eISSN:2075-4450

     More details

    Language:English   Publisher:Insects  

    Spermatogenesis is an important process in reproduction and is conserved across species, but in Bombyx mori, it shows peculiarities, such as the maintenance of spermatogonia by apical cells and fertilization by dimorphic spermatozoa. In this study, we attempted to characterize the genes expressed in the testis of B. mori, focusing on aspects of expression patterns and gene function by transcriptome comparisons between different tissues, internal testis regions, and Drosophila melanogaster. The transcriptome analysis of 12 tissues of B. mori, including those of testis, revealed the widespread gene expression of 20,962 genes and 1705 testis-specific genes. A comparative analysis of the stem region (SR) and differentiated regions (DR) of the testis revealed 4554 and 3980 specific-enriched genes, respectively. In addition, comparisons with D. melanogaster testis transcriptome revealed homologs of 1204 SR and 389 DR specific-enriched genes that were similarly expressed in equivalent regions of Drosophila testis. Moreover, gene ontology (GO) enrichment analysis was performed for SR-specific enriched genes and DR-specific enriched genes, and the GO terms of several biological processes were enriched, confirming previous findings. This study advances our understanding of spermatogenesis in B. mori and provides an important basis for future research, filling a knowledge gap between fly and mammalian studies.

    DOI: 10.3390/insects14080684

    Web of Science

    Scopus

    PubMed

  • The biological role of core 1?1-3galactosyltransferase (T-synthase) in mucin-type O-glycosylation in Silkworm,<i> Bombyx</i><i> mori</i>

    Morio, A; Lee, JM; Fujii, T; Mon, H; Masuda, A; Kakino, K; Xu, J; Banno, Y; Kusakabe, T

    INSECT BIOCHEMISTRY AND MOLECULAR BIOLOGY   156   103936   2023.5   ISSN:0965-1748 eISSN:1879-0240

     More details

    Language:English   Publisher:Insect Biochemistry and Molecular Biology  

    O-glycosylation of secreted and membrane-bound proteins is an important post-translational modification that affects recognition of cell surface receptors, protein folding, and stability. However, despite the importance of O-linked glycans, their biological functions have not yet been fully elucidated and the synthetic pathway of O-glycosylation has not been investigated in detail, especially in the silkworm. In this study, we aimed to investigate O-glycosylation in silkworms by analyzing the overall structural profiles of mucin-type O-glycans using LC–MS. We found GalNAc or GlcNAc monosaccharide and core 1 disaccharide (Galβ1-3-GalNAcα1-Ser/Thr) were major components of the O-glycan attached to secreted proteins produced in silkworms. Furthermore, we characterized the 1 b1,3-galactosyltransferase (T-synthase) required for synthesis of the core 1 structure, common to many animals. Five transcriptional variants and four protein isoforms were identified in silkworms, and the biological functions of these isoforms were investigated. We found that BmT-synthase isoforms 1 and 2 were localized in the Golgi apparatus in cultured BmN4 cells and functioned both in cultured cells and silkworms. Additionally, a specific functional domain of T-synthase, called the stem domain, was found to be essential for activity and is presumed to be needed for dimer formation and galactosyltransferase activity. Altogether, our results elucidated the O-glycan profile and function of T-synthase in the silkworm. Our findings allow the practical comprehension of O-glycosylation required for employing silkworms as a productive expression system.

    DOI: 10.1016/j.ibmb.2023.103936

    Web of Science

    Scopus

    PubMed

  • High yield production of norovirus GII.4 virus-like particles using silkworm pupae and evaluation of their protective immunogenicity

    Masuda, A; Lee, JM; Miyata, T; Sato, S; Masuda, A; Taniguchi, M; Fujita, R; Ushijima, H; Morimoto, K; Ebihara, T; Hino, M; Kakino, K; Mon, H; Kusakabe, T

    VACCINE   41 ( 3 )   766 - 777   2023.1   ISSN:0264-410X eISSN:1873-2518

     More details

    Language:English   Publisher:Vaccine  

    Noroviruses (NoVs) are one of the major causes of acute viral gastroenteritis in humans. Virus-like particles (VLPs) without genomes that mimic the capsid structure of viruses are promising vaccine candidates for the prevention of NoVs infection. To produce large amounts of recombinant protein, including VLPs, the silkworm-expression vector system (silkworm-BEVS) is an efficient and powerful tool. In this study, we constructed a recombinant baculovirus that expresses VP1 protein, the major structural protein of NoV GII.4. Expression analysis showed that the baculovirus-infected silkworm pupae expressed NoV VP1 protein more efficiently than silkworm larval fat bodies. We obtained about 4.9 mg of purified NoV VP1 protein from only five silkworm pupae. The purified VP1 protein was confirmed by dynamic light scattering and electron microscopy to form VLPs of approximately 40 nm in diameter. Antisera from mice immunized with the antigen blocked NoV VLPs binding to histo-blood group antigens of pig gastric mucin and also blocked NoV infection in intestinal epithelial cells derived from human induced pluripotent stem (iPS) cells. Our findings demonstrated that NoV VLP eliciting protective antibodies could be obtained in milligram quantities from a few silkworm pupae using the silkworm-BEVS.

    DOI: 10.1016/j.vaccine.2022.12.015

    Web of Science

    Scopus

    PubMed

  • Characterization of a Novel Heterochromatin Protein 1 Homolog "<i>HP1c</i>" in the Silkworm, <i>Bombyx mori</i>

    Hino, M; Tatsuke, T; Morio, A; Mon, H; Lee, JM; Masuda, A; Kakino, K; Tonooka, Y; Kusakabe, T

    INSECTS   13 ( 7 )   2022.7   ISSN:2075-4450 eISSN:2075-4450

     More details

    Language:English   Publisher:Insects  

    Heterochromatin protein 1 plays an important role in chromatin structure and gene expression regulation. Three HP1 genes have been found in Homo sapiens, and five HP1 genes have been reported in Drosophila melanogaster. On the other hand, in Bombyx mori, only two HP1 genes, BmHP1a and BmHP1b, were reported. In this research, we have reported the molecular and functional characterization of a novel Bombyx mori HP1 gene (BmHP1c), which had stronger transcriptional repression activity than BmHP1a. BmHP1a and BmHP1b is reported to form homo- and heterodimers, but in co-immunoprecipitation experiments, no homo- or hetero-dimer formation of BmHP1c with the other silkworm HP1s is detected. The intracellular localization of BmHP1c is not only in the nucleus but also in the cytoplasm like mammalian HP1γ. In contrast to human HP1a and b, all three BmHP1s were localized preferentially in the regions poorly stained with DAPI. Interestingly, the double knockdown of BmHP1a and b, but not BmHP1c with a or b, arrested the cell cycle at the G2/M phase. These results suggest that BmHP1c is not essential for cell progression and plays a different role than BmHP1a and BmHP1b.

    DOI: 10.3390/insects13070631

    Web of Science

    Scopus

    PubMed

  • Optimization of SARS-CoV-2 Spike Protein Expression in the Silkworm and Induction of Efficient Protective Immunity by Inoculation With Alum Adjuvants

    Masuda, A; Lee, JM; Miyata, T; Mon, H; Sato, K; Oyama, K; Sakurai, Y; Yasuda, J; Takahashi, D; Ueda, T; Kato, Y; Nishida, M; Karasaki, N; Kakino, K; Ebihara, T; Nagasato, T; Hino, M; Nakashima, A; Suzuki, K; Tonooka, Y; Tanaka, M; Moriyama, T; Nakatake, H; Fujita, R; Kusakabe, T

    FRONTIERS IN IMMUNOLOGY   12   803647   2022.1   ISSN:1664-3224

     More details

    Language:English   Publisher:Frontiers in Immunology  

    The newly emerged severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is causing a spread of coronavirus disease 2019 (COVID-19) globally. In order to end the COVID-19 pandemic, an effective vaccine against SARS-CoV-2 must be produced at low cost and disseminated worldwide. The spike (S) protein of coronaviruses plays a pivotal role in the infection to host cells. Therefore, targeting the S protein is one of the most rational approaches in developing vaccines and therapeutic agents. In this study, we optimized the expression of secreted trimerized S protein of SARS-CoV-2 using a silkworm-baculovirus expression vector system and evaluated its immunogenicity in mice. The results showed that the S protein forming the trimeric structure was the most stable when the chicken cartilage matrix protein was used as the trimeric motif and could be purified in large amounts from the serum of silkworm larvae. The purified S protein efficiently induced antigen-specific antibodies in mouse serum without adjuvant, but its ability to induce neutralizing antibodies was low. After examining several adjuvants, the use of Alum adjuvant was the most effective in inducing strong neutralizing antibody induction. We also examined the adjuvant effect of paramylon from Euglena gracilis when administered with the S protein. Our results highlight the effectiveness and suitable construct design of the S protein produced in silkworms for the subunit vaccine development against SARS-CoV-2.

    DOI: 10.3389/fimmu.2021.803647

    Web of Science

    Scopus

    PubMed

▼display all

Presentations

  • ヒトノロウイルスVLPを形成するVP1タンパク質の不溶性原因の解明と可溶性向上

    #鶴見祐土,#増田亮津,#李在萬,#門宏明,#藤田龍介,#日野真人,#藤井告,#日下部宜宏

    蚕糸・昆虫機能利用学術講演会  2024.3 

     More details

    Event date: 2024.3

    Language:Japanese   Presentation type:Oral presentation (general)  

    Venue:農林水産技術会議事務局筑波産学連携支援センター   Country:Japan  

  • 組換えタンパク質生産性向上の目指したランダム突然変異によるカイコバキュロウイルス変異体の獲得

    #渋谷美咲,#海老原健,#門宏明,#李在萬,#山口礼華,#増田亮津,#日野真人,#藤田龍介,#日下部宜宏

    蚕糸・昆虫機能利用学術講演会  2024.3 

     More details

    Event date: 2024.3

    Language:Japanese   Presentation type:Oral presentation (general)  

    Venue:農林水産技術会議事務局筑波産学連携支援センター   Country:Japan  

  • バキュロウイルス非必須遺伝子クラスター欠損株の構築とタンパク質発現解析

    #山口礼華,#海老原健,#門宏明,#李在萬,#藤田龍介,#増田亮津,#日野真人,#渋谷美咲,#日下部宜宏

    蚕糸・昆虫機能利用学術講演会  2024.3 

     More details

    Event date: 2024.3

    Language:Japanese   Presentation type:Oral presentation (general)  

    Venue:農林水産技術会議事務局筑波産学連携支援センター   Country:Japan  

  • カイコバキュロウイルス発現系を用いた高機能化多価ワクチン開発のための足場タンパク質ナノ粒子の探索

    #河合雄貴,#李在萬,#増田亮津,#日野真人,#藤田龍介,#日下部宜宏

    蚕糸・昆虫機能利用学術講演会  2024.3 

     More details

    Event date: 2024.3

    Language:Japanese   Presentation type:Oral presentation (general)  

    Venue:農林水産技術会議事務局筑波産学連携支援センター   Country:Japan  

  • カイコ-バキュロウイルス発現系を用いたナノ粒子化CSFワクチンの開発

    金子 大樹, 増田 亮津, 近藤 祐里奈, 和田 香里, 門 宏明, 日下部 宜宏, 李 在萬

    日本生物工学会大会講演要旨集  2024.8  (公社)日本生物工学会

     More details

    Language:Japanese  

  • カイコ-バキュロウイルス発現系を用いたナノ粒子化CSFワクチンの開発

    金子 大樹, 増田 亮津, 近藤 祐里奈, 和田 香里, 門 宏明, 日下部 宜宏, 李 在萬

    日本生物工学会大会講演要旨集  2024.8  (公社)日本生物工学会

     More details

    Language:Japanese  

▼display all

Professional Memberships

  • The Japanese Society Of Sericultural Science

  • The Molecular Biology Society of Japan

Research Projects

  • Development of a novel directed evolution method to improve recombinant protein productivity in insect factories

    Grant number:22K20581  2022 - 2023

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research  Grant-in-Aid for Research Activity start-up

    Masuda Akitsu

      More details

    Authorship:Principal investigator  Grant type:Scientific research funding

    In the silkworm-baculovirus expression system, which can efficiently produce recombinant proteins, it is known that there are some difficult-to-express proteins. Therefore, insect directed evolutionary tools that enable mutagenesis and selection of favourable mutants are needed to solve this problem. In this study, we attempted to develop a tool for introducing mutations into specific regions of the baculovirus genome and to identify baculovirus genes that affect viral proliferation, which is necessary for selecting desired mutants. The mutagenic enzyme was found to be expressed by the baculovirus in cultured insect cells, but no mutations in the target sequence were identified. In the identification of genes affecting viral proliferability, eight candidate genes were examined and several genes that could manipulate proliferability were identified.

    CiNii Research

  • ウイルス様粒子(VLP)表面への抗原提示法を用いた次世代型ワクチンの開発

    Grant number:19J21617 

    増田 亮津

      More details

    Grant type:Scientific research funding

    本研究は豚サーコウイルス2型(PCV2)のウイルス様粒子(VLP)の表面に豚流行性下痢病ウイルス(PEDV)のスパイク(S)抗原又は豚繁殖呼吸障害症候群ウイルス(PRRSV)のグリコプロテイン5(GP5)抗原をディスプレイすることで次世代型のワクチンを作り上げることである。そこで(1)ディスプレイ用抗原のカイコでの高効率生産、(2)九州大学のカイコ系統ライブラリーを利用したカイコの組換えタンパク質生産性の差異の原因探索、(3)タンパク質間のグルタミン残基とリジン残基を架橋する酵素である微生物由来トランスグルタミナーゼ (MTG)を用いた抗原提示VLPワクチン作製を課題として研究を進める。

    CiNii Research