2023 Fiscal Year Final Research Report
Structural analysis and developmental research of specific glycosidases for production of glycosides bearing biorecognition
| Project/Area Number |
18K05444
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Multi-year Fund |
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| Section | 一般 |
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| Review Section |
Basic Section 38030:Applied biochemistry-related
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| Research Institution | Osaka Metropolitan University (2022-2023)
Osaka City University (2018-2021) |
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Principal Investigator |
Ito Kazuo 大阪公立大学, 大学院理学研究科, 准教授 (20183171)
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| Project Period (FY) |
2018-04-01 – 2024-03-31
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| Keywords | 生体認識配糖体 / エンド-β-N-アセチルグルコサミニダーゼ / 立体構造 / ドメイン / マルチドメイン |
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| Outline of Final Research Achievements |
Crystal structure of recombinant endo-β-N-acetylglucosaminidase(Endo) HS resolved with 1.95Åresolution was revealed to be a novel multi-domain structure composed of 5 domains(D). The structure composed of D-I～III was similar to those of GHF85 enzymes, D-IV and V were characteristic of Endo HS. Four amino acid residues, N216, E218, Y252 and Y282 in D-I were revealed to compose catalytic site.
Crystal structure of recombinant Endo PMα, one of 3 isozymes of Endo PM from the isolate, Prevotella melaninogenica resolved with 2.1Å was composed of D-I～V from N-terminal. D-I was (α/β)8 barrel of catalytic D, D-II～V were βbarrels of different from each other. The structure composed of D-I～III was similar to those of GHF85 enzymes, D-IV and V were likely to be responsible for oligosaccharide recognition and sorting Endo PM to outer membrane, respectively. Small angle X-ray scattering analysis indicated that the structure in solution was different from the crystal structure.
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| Free Research Field |
酵素化学
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| Academic Significance and Societal Importance of the Research Achievements |
新規特異的糖鎖遊離酵素エンド-β-N-アセチルグルコサミニダーゼ(エンド)HSの立体構造が世界に先駆けて解明され、未知のドメインから構成される新規マルチドメイン構造が明らかとなった。既知糖鎖遊離酵素とは異なり、4アミノ酸残基が触媒部位を構成し、新規触媒機構を提唱する発見であった。
また、新分離菌P. melaninogenica 由来エンドPMのアイソザイム、エンドPMαの立体構造を初めて解明された。エンドPMαも未知ドメインで構成される新規マルチドメイン酵素で、各ドメインの構造上の特徴と機能が明らかとなった。また、本酵素の立体構造は、溶液中と結晶中では異なることが明らかとなった。
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