New High-Throughput Screening Assays for Biocatalysis

Abstract

High-throughput screening for catalysis is a critical technology in all experiments aimed at modifying or creating enzymes by directed evolution, as well as for biodiversity mining for new catalysts. We have developed a series of enzyme assays based on fluorogenic substrates and on fluorescent product sensors. These new assays offer the possibility to assay chemically non-activated functional groups within chiral molecules with unprecedented sensibility and selectivity. Assays are exemplified for alcohol dehydrogenases, aldolases, lipases and esterases, amidases and acylases, phosphatases, and epoxide hydrolases. The assays can also be used to isolate catalytic antibodies by screening libraries produced by immunization with transition-state analogs. These assays are suitable for microtiter plate and higher miniaturization formats.