title: Valine dehydrogenase from Streptomyces fradiae: purification and properties.
abstract: Valine dehydrogenase (VDH) was purified to homogeneity from cell-free extract of Streptomyces fradiae, which produces tylosin. The enzyme was purified 1508-fold in a 17.7% yield using a combination of hydrophobic chromatography and ion-exchange fast protein liquid chromatography. The Mr of the native enzyme was determined to be 218,000 and 215,000, by equilibrium ultracentrifugation and size-exclusion high-performance liquid chromatography, respectively. The enzyme is composed of 12 subunits of Mr 18,000. Using analytical isoelectric focusing the isoelectric point of VDH was found to be 4.7. Oxidative deamination of L-valine was optimal at pH 10.6. Reductive amination of 2-oxoisovalerate was optimal at pH 8.8. The Michaelis constants (Km) were 1 mM for L-valine and 0.029 mM for NAD+. Km values for reductive amination were 0.80 mM for 2-oxoisovalerate, 0.050 mM for NADH and 22 mM for NH4+.
authors.affiliation: Department of Fermentation Chemistry and Bioengineering, Prague Institute of Chemical Technology, Czechoslovakia.
authors.forename: A
authors.initials: A
authors.lastname: Vancura
authors.forename: I
authors.initials: I
authors.lastname: Vancurová
authors.forename: J
authors.initials: J
authors.lastname: Volc
authors.forename: S P
authors.initials: SP
authors.lastname: Fussey
authors.forename: M
authors.initials: M
authors.lastname: Flieger
authors.forename: J
authors.initials: J
authors.lastname: Neuzil
authors.forename: J
authors.initials: J
authors.lastname: Marsálek
authors.forename: V
authors.initials: V
authors.lastname: Bĕhal
mesh_terms: D000594:Amino Acid Oxidoreductases / Q000302:isolation & purification*; D002847:Chromatography, Agarose; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D003067:Coenzymes; D006863:Hydrogen-Ion Concentration; D008970:Molecular Weight; D013302:Streptomyces / Q000201:enzymology*; D013379:Substrate Specificity; D013696:Temperature; D050802:Valine Dehydrogenase (NADP+)
chemical_list: D003067:Coenzymes; D000594:Amino Acid Oxidoreductases; D050802:Valine Dehydrogenase (NADP+)