title: [Biochemical characterization and substrate profile of a highly enantioselective carbonyl reductase from Pichia pastoris].
abstract: Carbonyl reductases catalyze carbonyl compounds to chiral alcohols that are important building blocks in fine chemical industry. To study carbonyl reductase from Pichia pastoris GS115 (ppcr), we discovered a new gene (ppcr) encoding an NADPH-dependent carbonyl reductase by genomic data mining. It was amplified by PCR from the genomic DNA, and expressed in Escherichia coli BL21 (DE3). The recombinant protein was purified to homogeneity. The optimum temperature was 37 degrees C and the optimum pH of PPCR was 6.0. PPCR was stable below 45 degrees C. The Km and k(cat) value of the enzyme for ethyl 3-methyl-2-oxobutanoate were 9.48 mmol/L and 0.12 s, respectively. The enzyme had broad substrate specificity and high enantioselectivity. It catalyzed the reduction of aldehydes, a-ketoesters, beta-ketoesters and aryl ketones to give the corresponding alcohols with >97% ee with only a few exceptions, showing its application potential in the synthesis of chiral alcohols.
authors.affiliation: College of Bioengineering, Tianjin University of Science and Technology, Tianjin 300457, China.
authors.forename: Laiqiang
authors.initials: L
authors.lastname: Tian
authors.forename: Weidong
authors.initials: W
authors.lastname: Liu
authors.forename: Xi
authors.initials: X
authors.lastname: Chen
authors.forename: Jinhui
authors.initials: J
authors.lastname: Feng
authors.forename: Hongjiang
authors.initials: H
authors.lastname: Yang
authors.forename: Qiaqing
authors.initials: Q
authors.lastname: Wu
authors.forename: Dunming
authors.initials: D
authors.lastname: Zhu
authors.forename: Yanhe
authors.initials: Y
authors.lastname: Ma
mesh_terms: D000429:Alcohol Oxidoreductases / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D000595:Amino Acid Sequence; D001709:Biotechnology / Q000379:methods; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D010843:Pichia / Q000201:enzymology*; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D013237:Stereoisomerism; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D011994:Recombinant Proteins; D000429:Alcohol Oxidoreductases