2020 Fiscal Year Final Research Report
Engineered biosynthesis of new natural products for innovative drug discovery
Project Area | Creation of Complex Functional Molecules by Rational Redesign of Biosynthetic Machineries |
Project/Area Number |
16H06449
|
Research Category |
Grant-in-Aid for Scientific Research on Innovative Areas (Research in a proposed research area)
|
Allocation Type | Single-year Grants |
Review Section |
Science and Engineering
|
Research Institution | University of Shizuoka |
Principal Investigator |
|
Co-Investigator(Kenkyū-buntansha) |
植木 尚子 岡山大学, 資源植物科学研究所, 准教授 (50622023)
|
Project Period (FY) |
2016-06-30 – 2021-03-31
|
Keywords | 天然物 / 生合成 / 酵素反応機構 / 活性物質 / 遺伝子 / タンパク質構造解析 |
Outline of Final Research Achievements |
By structural characterizations, in vitro assays of CghA and its mutants and computational analyses, we determined how a Diels-Alderase (DAase) could evade product inhibition and how CghA exerts diastereoselectivity over the [4+2] cycloaddition reaction. CghA assumes a unique fold comprised of two flattened lipocalin-like beta-barrel domains. Structural analyses combined with in vitro assays of CghA mutants permitted rational engineering of CghA to favor the disfavored exo adduct formation, indicating that subtle tuning of the shape complementarity of the binding pocket to the reaction transition state is key to establishing desired diastereoselectivity in octalin-forming DAases. The CghA and 1 complex structure and computational analyses of the CghA catalyzed reaction show that CghA is optimized to bind higher-energy transition states than those involved in the nonenzymatic reaction pathways that give alternate isomers.
|
Free Research Field |
天然物化学
|
Academic Significance and Societal Importance of the Research Achievements |
Diels-Alderaseは一体どのようにして生成物阻害を回避しているのかといった学術上核心をなす疑問があった。我々は酵素CghAを発見し解析した結果、ジエンとジエノフィルを有する鎖状基質分子は酵素活性部位内で(E)-exo-enol体のコンフォメーションを保持され軌道の重なりと共に環化することが明らかとなった。しかしながら、天然物である生成物は(Z)-exo-enol体であることが既に知られていた。CghAは生成物(Z)-exo-enol体よりもエネルギー準位が高い(E)-exo-enol体を中間体とすることで生成物阻害を見事に回避し、極めて高い活性で環化生成物を放出することを突き止めた。
|