文献类型：期刊文献

英文题名：Practical two-step synthesis of enantiopure styrene oxide through an optimized chemoenzymatic approach

作者：Wu, Kai[1];Wang, Hualei[1];Chen, Lifeng[1];Fan, Haiyang[1];Zhao, Zhiqiang[1];Wei, Dongzhi[1]

机构：[1]East China Univ Sci & Technol, New World Inst Biotechnol, State Key Lab Bioreactor Engn, Shanghai 200237, Peoples R China

年份：2016

卷号：100

期号：20

起止页码：8757

外文期刊名：APPLIED MICROBIOLOGY AND BIOTECHNOLOGY

收录：;EI(收录号：20162202444748);WOS:【SCI-EXPANDED(收录号:WOS:000385128000012)】；

基金：This work was supported by the National Natural Science Foundation of China (No. 21406068/B060804), the Fundamental Research Funds for the Central Universities, and the National Basic Research Program of China (No. 2012CB721103).

语种：英文

外文关键词：Styrene oxide; Alcohol dehydrogenase; Enantioselectivity; Directed evolution

摘要：Enantiopure styrene oxide (SO) and its derivatives are important building blocks for chiral synthesis. In this study, we developed an attractive "1-pot, 2-step" chemoenzymatic approach for producing enantiopure SO with 100 % theoretical yield. This approach involved asymmetric reduction of a-chloroacetophenone by an alcohol dehydrogenase (ADH; step 1), followed by base-induced ring closure (epoxidation) of enantiopure 2-chloro-1-phenylethanol produced by the ADH (step 2). By-product formation during epoxidation was suppressed to <1 % by adding methyl tert-butyl ether (MTBE) as the second phase. Therefore, with this optimized approach, ADH from Lactobacillus kefir (LkDH) successfully produced 1 M (S)SO, with 99 % analytical yield and 97.8 % enantiomeric excess (ee). In the preparation of (R)-SO, a semi-rational strategy of active pocket iterative saturation mutagenesis (ISM) was successfully used to inverse the enantioselectivity of LkDH (muDH2, F147L/Y190P/A202F/M206H/V196L/S96D/K97V), which produced the opposite enantiomer (R)-2-chloro-1-phenylethanol. Through the optimized chemoenzymatic approach, muDH2 was successfully used to prepare 1 M (R)-SO, with 98.1 % ee and 99.0 % analytical yield. Our results indicated that this optimized chemoenzymatic approach could be used to produce both enantiomers of SO at concentrations as high as 120 g/L within 14 h, which is the highest concentration as far as we know. MuDH2 obtained through ISM also showed reversed enantioselectivity toward another 13 aromatic ketones, compared with wild-type (WT) LkDH. Furthermore, a molecular docking experiment demonstrated that muDH2 inverted the binding orientation of the substrate, which may be the reason for its inverse enantioselectivity.