Improved Production of (R)-1-phenyl-1,2-ethanediolby a Codon-ptimized R-specific Carbonyl Reductasefrom Candida parapsilosis in Escherichia coli

摘要

An R-specific carbonyl reductase from Candida parapsilosis (CprCR) catalyzesthe transformation of (R)-1-phenyl-1,2-ethanediol from 2-hydroxyacetophenone. The genercr coding CprCR contains a few codons rarely used by Escherichia coli. In order toimprove chiral alcohol production, three codon variants 24, aRCR, and mRCR of CprCRwere designed through truncation of 4–27 bp disorder sequence at the 5-terminus or/andadaption of nine rare codons. The effects of codon optimization on enzyme activity, proteinproduction, and otransformation were studied. Among these three types, the disordersequence-truncated and rare codon-adapted variant mRCR presents the highest enzymeactivity. When compared with CprCR, mRCR showed an increase of 35.6% in the totalactivity of cell-free extracts. The specific activity of mRCR presented similar increase in thecell-free extract with purified protein, which suggested that the codon optimization caused positive effect on protein productivity of variant enzyme. When microbial cellsconcentration was 30% (w/v), the molar conversion yield and enantiomeric excess of themRCR variant reached 86.4% and 93.6%, which were increased 36.5% and 15.8% thanthose of wild-type at a high substrate concentration of 5 g/L. The work will supply a newmethod for improving chiral alcohol preparation with codon engineeredmicroorganisms.