title: Upscale production of ethyl (S)-4-chloro-3-hydroxybutanoate by using carbonyl reductase coupled with glucose dehydrogenase in aqueous-organic solvent system.
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.
authors.affiliation: Institute of Bioengineering, Zhejiang University of Technology, Hangzhou, Zhejiang, 310014, People's Republic of China.
authors.forename: Zhi-Qiang
authors.initials: ZQ
authors.lastname: Liu
authors.forename: Jing-Jing
authors.initials: JJ
authors.lastname: Ye
authors.forename: Zhen-Yang
authors.initials: ZY
authors.lastname: Shen
authors.forename: Hua-Bin
authors.initials: HB
authors.lastname: Hong
authors.forename: Jian-Bo
authors.initials: JB
authors.lastname: Yan
authors.forename: Yi
authors.initials: Y
authors.lastname: Lin
authors.forename: Zheng-Xu
authors.initials: ZX
authors.lastname: Chen
authors.forename: Yu-Guo
authors.initials: YG
authors.lastname: Zheng
authors.forename: Yin-Chu
authors.initials: YC
authors.lastname: Shen
mesh_terms: D000429:Alcohol Oxidoreductases / Q000235:genetics / Q000378:metabolism*; D002087:Butyrates / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D042843:Glucose 1-Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D060847:Metabolic Engineering; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA; D012997:Solvents
chemical_list: D002087:Butyrates; D011994:Recombinant Proteins; D012997:Solvents; C121244:ethyl 4-chloro-3-hydroxybutanoate; D000429:Alcohol Oxidoreductases; D042843:Glucose 1-Dehydrogenase