Tuning Enzyme Activity for Nonaqueous Solvents: Engineering an Enantioselective "Michaelase" for Catalysis in High Concentrations of Ethanol.

ABSTRACT: Enzymes have evolved to function under aqueous conditions and may not exhibit features essential for biocatalytic application, such as the ability to function in high concentrations of an organic solvent. Consequently, protein engineering is often required to tune an enzyme for catalysis in non-aqueous solvents. In this study, we have used a collection of nearly all single mutants of 4-oxalocrotonate tautomerase, which promiscuously catalyzes synthetically useful Michael-type additions of acetaldehyde to various nitroolefins, to investigate the effect of each mutation on the ability of this enzyme to retain its "Michaelase" activity in elevated concentrations of ethanol. Examination of this mutability landscape allowed the identification of two hotspot positions, Ser30 and Ala33, at which

SUBMITTER: Guo C 

PROVIDER: S-EPMC7317446 | biostudies-literature | 2020 May

REPOSITORIES: biostudies-literature