Valine Dehydrogenase from Streptomyces hygroscopicus CH-7: Purification and Properties

• Published in: Actinomycetologica Vol. 13; no. 2; pp. 76 - 81
• Main Authors: Karadzic, I., Izrael-Tomaševic, A., Gojgic-Cvijovic, G., Vrvic, M. M.
• Format: Journal Article
• Language: English
• Published: The Society for Actinomycetes Japan 1999
• ISSN: 0914-5818; 1881-6371
• DOI: 10.3209/saj.13_76

Valine dehydrogenase (VDH) from Streptomyces hygroscopicus CH-7 was purified 327-fold with 21% yield, using ion exchange and hydrophobic chromatography. The enzyme had an Mr 31000 in denaturing conditions and an Mr 63000 in gel filtration chromatography, indicating that it is homo-dimer. The most preferable substrates are L-valine in deamination reaction and 2-ketoisovalerate in amination reaction. The enzyme requires NAD+ as a cofactor. The VDH from S. hygroscopicus CH-7 shows maximum activity at approximately pH 10.7 and 9.7 for deamination and amination reactions, respectively. The enzyme was significantly inhibited by p-chloromercuri-benzoate, Hg2+ and other metal ions, which suggests that the presence of SH- groups are necessary for the catalytic reaction. The apparent Michaelis constants for L-valine, NAD+ 2-ketoisovalerate, NADH and NH4+ were: 1.26 mM, 0.164 mM, 0.41 mM, 0.026 mM and 50.1 mM.