title: A novel D-mandelate dehydrogenase used in three-enzyme cascade reaction for highly efficient synthesis of non-natural chiral amino acids.
abstract: 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.
authors.affiliation: Laboratory of Biocatalysis and Synthetic Biotechnology, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China.
authors.forename: Chang-Wei
authors.initials: CW
authors.lastname: Fan
authors.affiliation: Laboratory of Biocatalysis and Synthetic Biotechnology, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China.
authors.forename: Guo-Chao
authors.initials: GC
authors.lastname: Xu
authors.affiliation: Laboratory of Biocatalysis and Synthetic Biotechnology, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China.
authors.forename: Bao-Di
authors.initials: BD
authors.lastname: Ma
authors.affiliation: Laboratory of Biocatalysis and Synthetic Biotechnology, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China.
authors.forename: Yun-Peng
authors.initials: YP
authors.lastname: Bai
authors.affiliation: Laboratory of Biocatalysis and Synthetic Biotechnology, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Laboratory of Biocatalysis and Synthetic Biotechnology, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China. Electronic address: jianhexu@ecust.edu.cn.
authors.forename: Jian-He
authors.initials: JH
authors.lastname: Xu
mesh_terms: D000429:Alcohol Oxidoreductases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D019149:Bioreactors; D005656:Fungal Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D005998:Glycine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D007778:Lactobacillus / Q000201:enzymology; D008333:Mandelic Acids / Q000378:metabolism; D016958:Pseudomonas putida / Q000201:enzymology; D019998:Racemases and Epimerases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D013237:Stereoisomerism
chemical_list: D005656:Fungal Proteins; D008333:Mandelic Acids; C008852:2-phenylglycine; D000429:Alcohol Oxidoreductases; C043630:D-mandelate dehydrogenase; D019998:Racemases and Epimerases; C021540:mandelate racemase; C037938:mandelic acid; D005998:Glycine
keywords: Asymmetric synthesis; Cascade reaction; D-Mandelate dehydrogenase; L-Phenylglycine; Process optimization