Budget Amount *help |
¥72,020,000 (Direct Cost: ¥55,400,000、Indirect Cost: ¥16,620,000)
Fiscal Year 2020: ¥14,040,000 (Direct Cost: ¥10,800,000、Indirect Cost: ¥3,240,000)
Fiscal Year 2019: ¥14,040,000 (Direct Cost: ¥10,800,000、Indirect Cost: ¥3,240,000)
Fiscal Year 2018: ¥14,040,000 (Direct Cost: ¥10,800,000、Indirect Cost: ¥3,240,000)
Fiscal Year 2017: ¥14,040,000 (Direct Cost: ¥10,800,000、Indirect Cost: ¥3,240,000)
Fiscal Year 2016: ¥15,860,000 (Direct Cost: ¥12,200,000、Indirect Cost: ¥3,660,000)
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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.
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