Significantly enhancing the biocatalytic synthesis of chiral alcohols by semi-rationally engineering an anti-Prelog carbonyl reductase from Acetobacter sp. CCTCC M209061

[Display omitted] •Chiral alcohols and their derivatives are important building blocks for the synthesis of high-valued chemicals.•Carbonyl reductases have been demonstrated to be valuable biocatalysts in the asymmetric reduction of carbonyl compounds.•For the activity of a carbonyl reductase AcCR to be improved, semi-rational engineering was performed.•Three AcCR variants were applied for chiral alcohols synthesis with excellent product yields and enantiomeric excess. Chiral alcohols and their derivatives are vital building blocks to synthesize pharmaceutical drugs and high-valued chemicals. Wild-type carbonyl reductase AcCR from Acetobacter sp. has ideal enantioselectivity toward 11 prochiral substrates (e.e.>99%) but poor activity. In this work, a semi-rational engineering was performed to enhance the activity of AcCR. Fortunately, three positive double-mutants (mut-E144A/G152 L, mut-G152 L/Y189 N, and mut-I147 V/G152 L) with specific activity 17–61 folds higher than that of enzyme without modified were achieved. Kinetic studies suggested that the catalytic efficiencies (kcat/Km) of these mutants were also well enhanced. Finally, these modified mut-AcCRs were successfully applied in asymmetric reductions of 11 structurally diverse prochiral substrates (200 mM) with excellent product yields (76.8%–99.1%) and enantiomeric excess (e.e.>99%), which provides an alternative strategy for efficient synthesis of chiral alcohols for pharmaceuticals industry with ideal yield and enantioselectivity.