title: Novel chiral tool, (R)-2-octanol dehydrogenase, from Pichia finlandica: purification, gene cloning, and application for optically active α-haloalcohols.
abstract: A novel enantioselective alcohol dehydrogenase, (R)-2-octanol dehydrogenase (PfODH), was discovered among methylotrophic microorganisms. The enzyme was purified from Pichia finlandica and characterized. The molecular mass of the enzyme was estimated to be 83,000 and 30,000 by gel filtration and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, respectively. The enzyme was an NAD(+)-dependent secondary alcohol dehydrogenase and showed a strict enantioselectivity, very broad substrate specificity, and high tolerance to SH reagents. A gene-encoding PfODH was cloned and sequenced. The gene consisted of 765 nucleotides, coding polypeptides of 254 amino acids. The gene was singly expressed and coexpressed together with a formate dehydrogenase as an NADH regenerator in an Escherichia coli. Ethyl (S)-4-chloro-3-hydroxybutanoate and (S)-2-chloro-1-phenylethanol were synthesized using a whole-cell biocatalyst in more than 99 % optical purity.
authors.affiliation: Green Product Development Center, R&D Management, Daicel Corporation, 1-1 Shinko-cho, Myoko, Niigata, 944-8550, Japan. h-yamamoto@jp.daicel.com
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Yamamoto
authors.forename: Masatake
authors.initials: M
authors.lastname: Kudoh
mesh_terms: D000429:Alcohol Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D015394:Molecular Structure; D008970:Molecular Weight; D009243:NAD / Q000378:metabolism; D000442:Octanols / Q000737:chemistry / Q000378:metabolism*; D010843:Pichia / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013379:Substrate Specificity
chemical_list: D005656:Fungal Proteins; D000442:Octanols; D009243:NAD; C067943:2-octanol; D000429:Alcohol Oxidoreductases; C021902:octanol dehydrogenase