Biosynthesis of (S)-4-chloro-3-hydroxybutanoate ethyl using Escherichia coli co-expressing a novel NADH-dependent carbonyl reductase and a glucose dehydrogenase

Bioresource Technology
Qi YeHanjie Ying

Abstract

A novel NADH-dependent carbonyl reductase (PsCR II) gene with an open reading frame of 855bp encoding 285 amino acids was cloned from Pichia stipitis. Analysis of the amino acid sequence of PsCR II revealed less than 55% identity to known reductases that produce (S)-4-chloro-3-hydroxybutanoates ethyl [(S)-CHBE]. When NADH was provided as an electron donor, Escherichia coli with pET-22b-PsCRII exhibited an activity of 15U/mg protein using 4-chloro-3-oxobutanoate ethyl (COBE) as a substrate. This activity was the highest ever reported for reductases, with the exception of PsCR I, which in our previous analysis required NADPH for catalysis. Biocatalysis of COBE to (S)-CHBE was investigated using E. coli with a polycistronic plasmid pET-BP II co-expressing PsCR II and a glucose dehydrogenase in a water/butyl acetate system for 24h. The transformants gave a molar yield of 91%, and an optical purity of the (S)-isomer of higher than 99% enantiomeric excess.

References

Apr 3, 2001·Current Opinion in Chemical Biology·J D Stewart
Nov 26, 2008·Chemico-biological Interactions·Bengt PerssonUdo Oppermann
Jul 14, 2009·Applied Microbiology and Biotechnology·Hiroyuki AsakoNobuya Itoh
Aug 5, 2009·Applied Microbiology and Biotechnology·Sang-Hyun Lee, Oh-Jin Park

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Citations

Mar 20, 2012·Bioresource Technology·Andrés IllanesGuillermo R Castro
Jul 14, 2012·Bioscience, Biotechnology, and Biochemistry·Mingdong AnLin Xu
Apr 9, 2017·Applied and Environmental Microbiology·Min LiJian-He Xu

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