Microbial Production of Ketoreductases: Development of a Novel High-Throughput Screening Method

Abstract

An assay method for detection of enantiospecific chiral alcohol was developed based on ketoreductase, enantio-selective alcohol oxidase and 2,4-dinitrophenyl hydrazine (DNPH) reagent. The assay method was developed to check the conversion of 1-Acetonapthone to either (S) or (R) specific 1-(1-napthyl) ethanol or its racemic mixture using ketoreductases. Further, estimation was done with the help of 2,4-DNPH method. The resulting orange coloured chromogen showed a maximum absorbance at 560 nm. The assay was performed in 96 well microtiter plates and had a linear detection range from 0.05 mM to 4 mM. The method is found to be suitable for the detection of large numbers of crude samples and screening of ketoreductase producing strains in high-throughput manner.