Biosynthesis of Ethyl (S)-4-Chloro-3-Hydroxybutanoate by NADH-Dependent Reductase from E. coli CCZU-Y10 Discovered by Genome Data Mining Using Mannitol as Cosubstrate

The reductase (PgCR) from recombinant Escherichia coli CCZU-Y10 displayed high reductase activity and excellent stereoselectivity for the reduction of ethyl 4-chloro-3-oxobutanoate (COBE) into ethyl (S)-4-chloro-3-hydroxybutanoate ((S)-CHBE). To efficiently synthesize (S)-CHBE (>99 % enantiomeric...

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Published in:Applied biochemistry and biotechnology 2014-08, Vol.173 (8), p.2042-2053
Main Authors: He, Yu-Cai, Yang, Zhen-Xing, Zhang, Dan-Ping, Tao, Zhi-Cheng, Chen, Chao, Chen, Yi-Tong, Guo, Fei, Xu, Jian-He, Huang, Lei, Chen, Rui-Jie, Ma, Xiao-Feng
Format: Article
Language:English
Subjects:
Acetoacetates - chemistry
Acetoacetates - metabolism
Biochemistry
Biological and medical sciences
Biosynthesis
Biotechnology
biotransformation
Chemistry
Chemistry and Materials Science
Data Mining
E coli
Enzyme Stability
Enzymes
Escherichia coli
Escherichia coli - enzymology
Escherichia coli - genetics
Escherichia coli Proteins - chemistry
Escherichia coli Proteins - genetics
Escherichia coli Proteins - metabolism
Fundamental and applied biological sciences. Psychology
genome
Genome, Bacterial
Genomes
Kinetics
mannitol
Mannitol - metabolism
NADH, NADPH Oxidoreductases - chemistry
NADH, NADPH Oxidoreductases - genetics
NADH, NADPH Oxidoreductases - metabolism
Stereoisomerism
Substrate Specificity
temperature
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Summary:The reductase (PgCR) from recombinant Escherichia coli CCZU-Y10 displayed high reductase activity and excellent stereoselectivity for the reduction of ethyl 4-chloro-3-oxobutanoate (COBE) into ethyl (S)-4-chloro-3-hydroxybutanoate ((S)-CHBE). To efficiently synthesize (S)-CHBE (>99 % enantiomeric excess (ee)), the highly stereoselective bioreduction of COBE into (S)-CHBE with the whole cells of E. coli CCZU-Y10 was successfully demonstrated in a dibutyl phthalate-water biphasic system. The appropriate ratio of the organic phase to water phase was 1:1 (v/v). The optimum reaction temperature, reaction pH, cosubstrate, NAD⁺, and cell dosage of the biotransformation of 100 mM COBE in this biphasic system were 30 °C, 7.0, mannitol (2.5 mmol/mmol COBE), 0.1 μmol/(mmol COBE), and 0.1 g (wet weight)/mL, respectively. Moreover, COBE at a high concentration of (1,000 mM) could be asymmetrically reduced to (S)-CHBE in a high yield (99.0 %) and high enantiometric excess value (>99 % ee). Significantly, E. coli CCZU-Y10 shows high potential in the industrial production of (S)-CHBE (>99 % ee).
ISSN:0273-2289
1559-0291
DOI:10.1007/s12010-014-1001-4