Bienzymatic Dynamic Kinetic Resolution of Secondary Alcohols by Esterification/Racemization in Water

Abstract

Dynamic kinetic resolution (DKR) is a key method used to prepare optically pure compounds in 100% theoretical yield starting from racemic substrates by combining the interconversion of substrate enantiomers with an enantioselective transformation. Various chemoenzymatic DKR approaches have been developed to deracemize secondary alcohols, typically requiring an organic solvent to facilitate enantioselective acylation, primarily catalyzed by lipases, alongside racemization mediated by an achiral, non-enzymatic catalyst. Achieving both steps in an aqueous solution remained elusive. Here, we report a DKR of racemic sec-alcohols in an aqueous solution requiring only two biocatalysts. The first key to success was to achieve fast racemization in a buffer employing a non-stereoselective variant of alcohol dehydrogenase (Lk-ADH-Prince) via a hydrogen-borrowing oxidation-reduction sequence. Engineered variants of the acyltransferase from Mycobacterium smegmatis (MsAcT) enabled enantioselective acyl transfer in water. Besides the appropriate choice of the enzymes, identifying a suitable acyl donor was a second key to the success. The DKR was successfully demonstrated using (R)-selective MsAcT variants for a broad range of racemic (hetero)benzylic alcohols using 2,2,2-trifluoroethyl acetate as the acyl donor, yielding (R)-acetates with up to >99% conv. and high-to-excellent optical purity (83–99.9% ee). The (S)-acetates were accessible using a stereocomplementary (S)-selective MsAcT variant.