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Practical two-step synthesis of enantiopure styrene oxide through an optimized chemoenzymatic approach

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Abstract

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 α-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.

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Acknowledgments

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).

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    1. State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai, 200237, People’s Republic of China

      Kai Wu, Hualei Wang, Lifeng Chen, Haiyang Fan, Zhiqiang Zhao & Dongzhi Wei

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    1. Kai Wu
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    2. Hualei Wang
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    Correspondence to Hualei Wang or Dongzhi Wei.

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    Kai Wu and Hualei Wang contributed equally to this work.

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    Wu, K., Wang, H., Chen, L. et al. Practical two-step synthesis of enantiopure styrene oxide through an optimized chemoenzymatic approach. Appl Microbiol Biotechnol 100, 8757–8767 (2016). https://doi.org/10.1007/s00253-016-7631-4

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