title: Cloning and overexpression of ketopantoic acid reductase gene from Stenotrophomonas maltophilia and its application to stereospecific production of D-pantoic acid.
abstract: Ketopantoic acid (KPA) reductase catalyzes the stereospecific reduction of ketopantoic acid to D-pantoic acid. Based on the N-terminal amino acid sequence of KPA reductase from Stenotrophomonas maltophilia 845, the KPA reductase gene was cloned from S. maltophilia NBRC14161 and sequenced. This gene contains an open reading frame of 777 bp encoding 258 amino acid residues, and the deduced amino acid sequence showed high similarity to the SDR superfamily proteins. An expression vector, pETSmKPR, containing the full KPA reductase gene was constructed and introduced into Escherichia coli BL21 (DE3) to overexpress the enzyme. Bioreduction of KPA using E. coli transformant cells coexpressing KPA reductase together with cofactor regeneration enzyme gene was also performed. The conversion yield of KPA to D-pantoic acid reached over 88% with a substrate concentration up to 1.17 M.
authors.affiliation: Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kyoto, Japan.
authors.forename: Dayong
authors.initials: D
authors.lastname: Si
authors.forename: Nobuyuki
authors.initials: N
authors.lastname: Urano
authors.forename: Sakayu
authors.initials: S
authors.lastname: Shimizu
authors.forename: Michihiko
authors.initials: M
authors.lastname: Kataoka
mesh_terms: D000429:Alcohol Oxidoreductases / Q000235:genetics* / Q000378:metabolism*; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D006885:Hydroxybutyrates / Q000378:metabolism*; D007651:Keto Acids / Q000378:metabolism*; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D020615:Stenotrophomonas maltophilia / Q000201:enzymology* / Q000235:genetics
chemical_list: D006885:Hydroxybutyrates; D007651:Keto Acids; D011994:Recombinant Proteins; C016031:pantoic acid; C015963:2-ketopentanoic acid; D000429:Alcohol Oxidoreductases; C021934:2-dehydropantoate 2-reductase