A novel D-mandelate dehydrogenase used in three-enzyme cascade reaction for highly efficient synthesis of non-natural chiral amino acids.

J Biotechnol

Laboratory of Biocatalysis and Synthetic Biotechnology, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China. Electronic address:

Published: February 2015

A novel NAD(+)-dependent D-mandelate dehydrogenase was identified from Lactobacillus brevis (LbDMDH). After purified to homogeneity, the optimum pH and temperature for oxidation of D-mandelate were pH 10.0 and 40 °C, and the Km and kcat were 1.1 mM and 355 s(-1) respectively. Employing the LbDMDH together with a mandelate racemase from Pseudomonas putida and a leucine dehydrogenase (EsLeuDH) from Exiguobacterium sibiricum, we established a three-step one-pot domino reaction system for preparing chiral L-phenylglycine from racemic mandelic acid with internal cofactor recycling. Under the optimum conditions, 30.4 g rac-mandelic acid (0.2 M) at 1L scale had been converted into chiral L-phenylglycine, with 96.4% conversion, 86.5% isolation yield, >99% eep and 50.4 gL(-1)d(-1) space-time yield.

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http://dx.doi.org/10.1016/j.jbiotec.2014.10.026DOI Listing

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