Expression system of CotA-laccase for directed evolution and high-throughput screenings for the oxidation of high-redox potential dyes

Laccases are useful biocatalysts for many diverse biotechnological applications. In this study wehave established efficient and reliable expression systems and high-throughput screenings for therecombinant CotA-laccase from Bacillus subtilis. The expression levels of cotA-laccase were comparedin five different Escherichia coli host strains growing in 96-well microtiter plates under differentculture conditions. Lower coefficients of variance (around 15%) were achieved using crude celllysates of BL21 and KRX host strains growing under microaerobic conditions. Reproducible highthroughputscreenings for the decolorization of high redox potential azo and anthraquinonic dyeswere developed and optimized for identification of variants with increased redox potential. The enzymaticassays developed were tested for the screening of one mutant library from CotA-laccasecreated by error-prone PCR.