| Project/Area Number |
17K07708
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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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| Research Field |
Applied microbiology
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| Research Institution | Yamagata University |
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Principal Investigator |
YANO Shigekazu 山形大学, 大学院理工学研究科, 准教授 (50411228)
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| Co-Investigator(Kenkyū-buntansha) |
今野 博行 山形大学, 大学院理工学研究科, 教授 (50325247)
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| Project Period (FY) |
2017-04-01 – 2020-03-31
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| Project Status |
Completed (Fiscal Year 2019)
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| Budget Amount *help |
¥4,810,000 (Direct Cost: ¥3,700,000、Indirect Cost: ¥1,110,000)
Fiscal Year 2019: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000)
Fiscal Year 2018: ¥1,820,000 (Direct Cost: ¥1,400,000、Indirect Cost: ¥420,000)
Fiscal Year 2017: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
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| Keywords | α-1,3-グルカナーゼ / 転移反応 / ニゲロオリゴ糖 / 結晶構造解析 |
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| Outline of Final Research Achievements |
α-1,3-Glucanase Agl-KA from Bacillus circulans KA-304 hydrolyzes the α-1,3-glucoside linkages in α-1,3-glucan. In this study, we investigated transglycosylation activity of Agl-KA under a low water content. Lyophilized Agl-KA and nigero-oligosaccharides were stored at 80% humidity. After incubation, transglycosylation activity was confirmed, and the optimum pH of the transferring reaction was 10.
We also analyzed molecular structure of the catalytic domain of Agl-KA. The structure was determined using X-ray crystallography. The catalytic unit consists of a complex structure of two tandemly connected regions, the N-terminal galactose-binding-like region and the C-terminal right-handed β-helix region. Biochemical assays have found that Asp1067, Asp1090, and Asp1091 are important for catalysis, and these residues indeed locate to the putative substrate binding cleft, which forms a closed end and explains the product specificity.
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| Academic Significance and Societal Importance of the Research Achievements |
本研究を行うことで、新規のニゲロオリゴ糖生産法を示した。低含水状態、あるいは高濃度基質存在下でα-1,3-グルカナーゼの転移反応が進行したことから、様々な加水分解酵素でも同様の条件下で転移反応が進行する可能性がある。この技術は、新規のオリゴ糖、配糖体やペプチドの合成を可能にするので、食品業界に広く普及する。セミドライでの転移反応は、乾燥食品中でも起きている可能性があり、乾燥食品の味の秘密を解き明かすのに役立つと考えている。
また、本研究では、α-1,3-グルカナーゼ触媒ドメインの結晶構造を明らかにした。本結果をもとに、産業利用に適した改変型α-1,3-グルカナーゼの作製も期待できる。
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