5X8H

Crystal structure of the ketone reductase ChKRED20 from the genome of Chryseobacterium sp. CA49

5X8H の概要

• エントリーDOI: 10.2210/pdb5x8h/pdb
• 分子名称: Short-chain dehydrogenase reductase (2 entities in total)
• 機能のキーワード: oxidoreductase
• 由来する生物種: Chryseobacterium sp. CA49
• タンパク質・核酸の鎖数: 4
• 化学式量合計: 105035.19

構造登録者

Zhao, F.J.,Jin, Y.,Liu, Z.C.,Wang, G.G.,Wu, Z.L. (登録日: 2017-03-02, 公開日: 2017-11-08, 最終更新日: 2023-11-22)

主引用文献

Zhao, F.J.,Jin, Y.,Liu, Z.,Guo, C.,Li, T.B.,Li, Z.Y.,Wang, G.,Wu, Z.L.
Crystal structure and iterative saturation mutagenesis of ChKRED20 for expanded catalytic scope
Appl. Microbiol. Biotechnol., 101:8395-8404, 2017

PubMed Abstract: ChKRED20 is an efficient and robust anti-Prelog ketoreductase that can catalyze the reduction of ketones to chiral alcohols as pharmaceutical intermediates with great industrial potential. To overcome its limitation on the bioreduction of ortho-substituted acetophenone derivatives, the X-ray crystal structure of the apo-enzyme of ChKRED20 was determined at a resolution of 1.85 Å and applied to the molecular modeling and reshaping of the catalytic cavity via three rounds of iterative saturation mutagenesis together with alanine scanning and recombination. The mutant Mut3B was achieved with expanded catalytic scope that covered all the nine substrates tested as compared with two substrates for the wild type. It exhibited 13-20-fold elevated k /K values relative to the wild type or to the first gain-of-activity mutant, while retaining excellent stereoselectivity toward seven of the substrates (98-> 99% ee). Another mutant 29G10 displayed complementary selectivity for eight of the ortho-substituted acetophenone derivatives, with six of them delivering excellent stereoselectivity (90-99% ee). Its k /K value toward 1-(2-fluorophenyl)ethanone was 5.6-fold of the wild type. The application of Mut3B in elevated substrate concentrations of 50-100 g/l was demonstrated in 50-ml reactions, achieving 75-> 99% conversion and > 99% ee.

PubMed: 29067484
DOI: 10.1007/s00253-017-8556-2

実験手法

X-RAY DIFFRACTION (1.85 Å)