title: 3-Hydroxyisobutyrate dehydrogenase from Pseudomonas putida E23: purification and characterization.
abstract: The NAD(+)-dependent 3-hydroxyisobutyrate dehydrogenase [EC 1.1.1.31] was purified to homogeneity from Pseudomonas putida E23. The enzyme was a tetramer (molecular mass, 120 kDa) consisted of identical subunits (molecular mass, 30 kDa). The enzyme was specific for NAD+ (Km, 0.44 mM). The maximal activity was obtained at about pH 10. The enzyme was specific for the L-isomer of 3-hydroxyisobutyrate. In addition to L-3-hydroxyisobutyrate, L-serine, 2-methyl-DL-serine, and 3-hydroxypropionate were substrates. The Km for L-3-hydroxyisobutyrate, L-serine, 2-methyl-DL-serine, and 3-hydroxypropionate were 0.12, 18, 44, and 83 mM, respectively. The enzyme was inhibited by p-chloromercuribenzoate, HgCl2, and AgNO3, but not by EDTA, alpha,alpha'-dipyridyl, and o-phenanthroline. The N-terminal 26 amino acid sequence was compared with the sequences deduced from the enzyme genes of rat liver and Pseudomonas aeruginosa.
authors.affiliation: Department of Bioresources Sciences, Kochi University, Japan.
authors.forename: E K
authors.initials: EK
authors.lastname: Chowdhury
authors.forename: S
authors.initials: S
authors.lastname: Nagata
authors.forename: H
authors.initials: H
authors.lastname: Misono
mesh_terms: D000429:Alcohol Oxidoreductases / Q000032:analysis* / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D000595:Amino Acid Sequence; D000818:Animals; D003067:Coenzymes / Q000378:metabolism; D003470:Culture Media; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016958:Pseudomonas putida / Q000201:enzymology*; D051381:Rats; D012988:Soil Microbiology; D013056:Spectrophotometry, Ultraviolet; D013379:Substrate Specificity
chemical_list: D003067:Coenzymes; D003470:Culture Media; D000429:Alcohol Oxidoreductases; C021398:3-hydroxyisobutyrate dehydrogenase