title: Cloning, overexpression, and characterization of a high enantioselective nitrilase from Sphingomonas wittichii RW1 for asymmetric synthesis of (R)-phenylglycine.
abstract: In this study, a high (R)-enantioselective nitrilase gene from Sphingomonas wittichii RW1 was cloned and overexpressed in Escherichia coli BL21 (DE3). The recombinant nitrilase was purified to homogeneity with a molecular weight of 40 kDa. The pH and temperature optima were shown to be pH 8.0 and 40 °C, respectively. The purified nitrilase was most active toward succinonitrile, approximately 30-fold higher than that for phenylglycinonitrile. Using the E. coli BL21/ReSWRW1 whole cells as biocatalysts, the kinetic resolution for asymmetric synthesis of (R)-phenylglycine was investigated at pH 6.0. A yield of 46 % was obtained with 95 % enantiomeric excess (ee), which made it a promising biocatalyst for synthesis of (R)-phenylglycine.
authors.affiliation: State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai, 200237, People's Republic of China.
authors.forename: Jian
authors.initials: J
authors.lastname: Qiu
authors.forename: Er-Zheng
authors.initials: EZ
authors.lastname: Su
authors.forename: Hua-Lei
authors.initials: HL
authors.lastname: Wang
authors.forename: Wen-Wen
authors.initials: WW
authors.lastname: Cai
authors.forename: Wei
authors.initials: W
authors.lastname: Wang
authors.forename: Dong-Zhi
authors.initials: DZ
authors.lastname: Wei
mesh_terms: D000595:Amino Acid Sequence; D000619:Aminohydrolases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005998:Glycine / Q000031:analogs & derivatives* / Q000096:biosynthesis / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008670:Metals / Q000494:pharmacology; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D017421:Sequence Analysis; D020578:Sphingomonas / Q000201:enzymology* / Q000235:genetics*; D013237:Stereoisomerism; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D008670:Metals; D011994:Recombinant Proteins; C008852:2-phenylglycine; D000619:Aminohydrolases; C021867:nitrilase; D005998:Glycine