Multi-enzymatic Systems for the Production of Chiral Compounds

Abstract

Two multi-enzymatic systems for the production of optically active compounds are discussed. One is a system for deracemization via enzymatic stereoinversion useful for chiral 1,2-diol synthesis, which involves stereoselective oxidation and stereoselective reduction using an alcohol dehydrogenase and reductase with opposite stereospecificities. The recycling of NAD⁺ was carried out by a water-forming NADH oxidase. The recycling of NADPH was carried out by an NADPH-dependent glucose dehydrogenase. The recombinant Escherichia coli capable of overproducing the four enzymes was constructed for purposes of generating catalysts useful for the deracemization of racemic diols. Optically active 3-chloro-1,2-propanediol and (S)-1,2-pentanediol could be prepared by using the recombinant E. coli cells in a one-pot reaction with high yield. Second is a transamination system for chiral secondary amine synthesis. Two novel bacterial ω-transaminases capable of catalyzing a stereoselective transamination between a ketone and amine were discovered by enrichment culturing. An (R)-specific ω-transaminase was isolated from a microorganism and characterized, and the relevant gene was cloned. Addition of lactate dehydrogenase into the transaminase reaction mixture was effective at increasing the yield by ensuring the reversibility of the reaction. A recombinant E. coli system capable of overproducing the ω-transaminase, lactate dehydrogenase, and glucose dehydrogenase used for regenerating NADH was constructed. By using the recombinant enzyme systems, various kinds of useful optically active amines could be prepared in good yields and high optical purity without using high-pressure and high-temperature conditions.