High-Throughput Screening for a Biocatalytic Asymmetric Ketone Reduction

Significance

The authors report a high-throughput, 96-well microtiterplate-format screening method for determining both the activity and the enantioselectivity of a biocatalytic asymmetric ketone reduction. Reaction of acetophenone (1) with NADPH in the presence of a ketoreductase (KRED) gives 1-phenylethanols (R)- and (S)-2. The rate is determined by monitoring the consumption of NADPH through the decrease in absorbance at 340 nm (activity). An (R)-selective alcohol oxidase catalyzes the oxidation of alcohol (R)-2 back to ketone 1 with concomitant production of H₂O₂. The amount of H₂O₂, and hence (R)-alcohol is monitored at 400 nm through a colorimetric HRP/ABTS assay (enantioselectivity). Screening of 17 different KREDs revealed a good correlation between this new assay and the conventional HPLC analysis for both activity and enantioselectivity.