Mechanistic study for stereochemical control of microbial reduction of α-keto esters in an organic solvent

Abstract

To elucidate the mechanism for stereochemical control of yeast reduction of α-keto esters in organic media, seven enzymes responsible for the reduction have been isolated from bakers' yeast and kinetic parameters for enzymatic reductions have been measured. In yeast reduction of ethyl 3-methyl-2-oxobutanoate ( 1f), enantiomeric excess in the produced ( R)-hydroxy ester increases when an organic solvent is used as the reaction medium in place of water. Difference in K m of the enzymes contributes largely to the stereochemistry of reduction by whole yeast cell. Four enzymes contribute to catalytic reduction of 1f. K m of the ( R)-producing enzyme has been found to be the lowest among those of four enzymes. Stereochemical course of the reduction shifts toward the ( R)-product by lowering the substrate concentrations, because the ( R)-producing enzyme is most active among the enzymes under diluted conditions. In yeast reduction of ethyl 2-oxohexanoate ( 1d), the corresponding ( S)-hydroxy ester is obtained in water, whereas the antipode is given in benzene. Five enzymes participate to the reduction of 1d and the ( R)-producing enzymes have smaller K ms than those of the ( S)-producing enzymes. When the reaction is run in benzene, however, the produced α-hydroxy ester does not undergo further decomposition. The inhibition of enzymatic decomposition in an organic solvent is also accounted for by low concentration of α-hydroxy ester in aqueous phase surrounding the bakers' yeast.