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[Construction of an enzyme-coupled system consisting of (R)- and (S)-specific carbonyl reductases for one-step preparation of (S)-1-phenyl-1,2-ethanediol].

Wei sheng wu xue bao = Acta microbiologica Sinica (2009-05-19)
Rongzhen Zhang, Yan Xu, Ying Sun, Yawei Geng
ABSTRACT

To prepare (S)-1-phenyl-1,2-ethanediol by a one-step method, we constructed an enzyme coupled system consisting of (R)- and (S)-specific carbonyl reductases in Escherichia coli. The genes coding (R)- and (S)-specific carbonyl reductases from Candida parapsilosis were inserted into a co-expression vector pETDuet-1. The positive plasmid was transformed into codon optimized E. coli Rosetta and an enzyme-coupled recombinant strain E. coli Rosetta / pETDuet-rcr-fdh was constructed. When the OD600 value of the culture reached 0.6-0.8, IPTG with a final concentration of 1 mmol/L was added to induce both target proteins at 30 degrees C for 10 h. SDS-PAGE analysis showed that two target enzymes were expressed simultaneously with the relative molecular weights of 37 kDa and 30 kDa. When 5 mol/L Zn2+ was added into a phosphate buffer (pH7.0), the biotransformation results showed that the product (S)-1-phenyl-1,2-ethanediol was produced with the optical purity of 91.3% enantiomeric excess and a yield of 75.9% by E. coli Rosetta/pETDuet-rcr-scr. The functions of two redox enzymes were integrated by molecular recombinant technique and a one-step method for preparation of (S)-1-phenyl-1,2-ethanediol was developed through an enzyme-coupled system. The method may supply a new procedure in chiral alcohols preparation.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
1-Phenyl-1,2-ethanediol, 97%
Sigma-Aldrich
(S)-(+)-1-Phenyl-1,2-ethanediol, 99%
Sigma-Aldrich
(R)-(−)-1-Phenyl-1,2-ethanediol, 99%