A Novel Whole-Cell Biocatalyst with NAD(+) Regeneration for Production of Chiral Chemicals

BACKGROUND: The high costs of pyridine nucleotide cofactors have limited the applications of NAD(P)-dependent oxidoreductases on an industrial scale. Although NAD(P)H regeneration systems have been widely studied, NAD(P)(+) regeneration, which is required in reactions where the oxidized form of the cofactor is used, has been less well explored, particularly in whole-cell biocatalytic processes. METHODOLOGY/PRINCIPAL FINDINGS: Simultaneous overexpression of an NAD(+) dependent enzyme and an NAD(+) regenerating enzyme (H(2)O producing NADH oxidase from Lactobacillus brevis) in a whole-cell biocatalyst was studied for application in the NAD(+)-dependent oxidation system. The whole-cell biocatalyst with (2R,3R)-2,3-butanediol dehydrogenase as the catalyzing enzyme was used to produce (3R)-acetoin, (3S)-acetoin and (2S,3S)-2,3-butanediol. CONCLUSIONS/SIGNIFICANCE: A recombinant strain, in which an NAD(+) regeneration enzyme was coexpressed, displayed significantly higher biocatalytic efficiency in terms of the production of chiral acetoin and (2S,3S)-2,3-butanediol. The application of this coexpression system to the production of other chiral chemicals could be extended by using different NAD(P)-dependent dehydrogenases that require NAD(P)(+) for catalysis.