Abstract
(S)-4-chloro-3-hydroxybutanoate ((S)-CHBE) is an important chiral intermediate to synthesize the side chain of cholesterol-lowering drug atorvastatin. To biosynthesize the (S)-CHBE, a recombinant Escherichia coli harboring the carbonyl reductase and glucose dehydrogenase was successfully constructed. The recombinant E. coli was cultured in a 500-L fermentor; after induction and expression, the enzyme activity and cell biomass were increased to 23,661.65 U/L and 13.90 g DCW/L which was 3.24 and 2.60-folds compared with those in the 50 L fermentor. The biocatalytic process for the synthesis of (S)-CHBE in an aqueous-organic solvent system was constructed and optimized with a substrate fed-batch strategy. The ethyl 4-chloro-3-oxobutanoate concentration reached to 1.7 M, and the (S)-CHBE with yield of 97.2 % and enantiomeric excess (e.e.) of 99 % was obtained after 4-h reaction in a 50-L reactor. In this study, the space-time yield and space-time yield per gram of biomass (dry cell weight, DCW) were 413.17 mM/h and 27.55 mM/h/g DCW for (S)-CHBE production, respectively, which were the highest values as compared to previous reports. Finally, (S)-CHBE was extracted from the reaction mixture with 82 % of yield and 95 % of purity. This study paved the foundation for the upscale production of (S)-CHBE by biocatalysis method.
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Acknowledgments
This work was supported by the National Basic Research Program of China (973 Program) (No. 2011CB710800), Natural Science Foundation of Zhejiang Province (No. R3110155) and Qianjiang Talent Project of Zhejiang Province.
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Liu, ZQ., Ye, JJ., Shen, ZY. et al. Upscale production of ethyl (S)-4-chloro-3-hydroxybutanoate by using carbonyl reductase coupled with glucose dehydrogenase in aqueous-organic solvent system. Appl Microbiol Biotechnol 99, 2119–2129 (2015). https://doi.org/10.1007/s00253-014-6245-y
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DOI: https://doi.org/10.1007/s00253-014-6245-y