ABSTRACT: (2S,3S)-2,3-Butanediol ((2S,3S)-2,3-BD) is a potentially valuable liquid fuel and an excellent building block in asymmetric synthesis. In this study, cofactor engineering was applied to improve the efficiency of (2S,3S)-2,3-BD production and simplify the product purification. Two NADH regeneration enzymes, glucose dehydrogenase and formate dehydrogenase (FDH), were introduced into Escherichia coli with 2,3-BD dehydrogenase, respectively. Introduction of FDH resulted in higher (2S,3S)-2,3-BD concentration, productivity and yield from diacetyl, and large increase in the intracellular NADH concentration. In fed-batch bioconversion, the final titer, productivity and yield of (2S,3S)-2,3-BD on diacetyl reached 31.7 g/L, 2.3 g/(L·h) and 89.8%, the highest level of (2S,3S)-2,3-BD production thus far. Moreover, cosubstrate formate was almost totally converted to carbon dioxide and no organic acids were produced. The biocatalytic process presented should be a promising route for biotechnological production of NADH-dependent microbial metabolites.
