title: Kinetic study of thermostable L-threonine dehydrogenase from an archaeon Pyrococcus horikoshii.
abstract: In the genome data base of the hyperthermophilic archaeon Pyrococcus horikoshii, an open reading frame with sequence homology to a gene encoding alcohol dehydrogenase was found. It was demonstrated that the encoded enzyme was a thermostable L-threonine dehydrogenase which can oxidize the hydroxy alkyl residue of L-threonine associated with the reduction of NAD+ or NADP+. This enzyme is a member of the zinc-containing L-threonine dehydrogenase family. One enzyme molecule contained one zinc atom, and this metal was considered to contribute to the hyperthermostablility of the enzyme. The reaction of the enzyme proceeded via a sequential mechanism. The Michaelis constants (Km) for L-threonine and NAD+ were 0.013 and 0.010 mM, respectively, and the maximum reaction rate (Vmax) was 1.75 mmol NADH formed/min/mg-protein at 65 degrees C. The Km values for both L-threonine and NADP+ were larger than those for L-threonine and NAD+ with a similar Vmax value. These results indicate that the enzyme has lower affinity to NADP+ than to NAD+, and the binding affinity for L-threonine depends on the coenzymes.
authors.affiliation: Research Institute for Cell Engineering, National Institute of Advanced Industrial Science and Technology, 1-8-31 Midorigaoka, Ikeda, Osaka 563-8577, Japan.
authors.forename: Noriko
authors.initials: N
authors.lastname: Higashi
authors.forename: Harumi
authors.initials: H
authors.lastname: Fukada
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Ishikawa
mesh_terms: D000429:Alcohol Oxidoreductases / Q000032:analysis* / Q000737:chemistry* / Q000235:genetics; D000595:Amino Acid Sequence; D004789:Enzyme Activation; D004795:Enzyme Stability; D007700:Kinetics; D008969:Molecular Sequence Data; D043902:Pyrococcus horikoshii / Q000201:enzymology* / Q000235:genetics; D011994:Recombinant Proteins / Q000032:analysis / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D013696:Temperature
chemical_list: D011994:Recombinant Proteins; D000429:Alcohol Oxidoreductases; C023159:L-threonine 3-dehydrogenase