Purification and Some Properties of Glutaminase from Pseudomonas nitroreducens IFO 12694

• Published in: Bioscience, biotechnology, and biochemistry biotechnology, and biochemistry, 1996, Vol.60 (7), p.1160-1164
• Main Authors: Takashi, Tachiki, Takeshi, Yamada, Masashi, Ueda, Yasuki, Naemura, Nobuo, Imamura, Yu-ichi, Hamada, Ju-ichi, Shiode
• Format: Article
• Language: English
• Subjects: Bacteriology; Biological and medical sciences; Biology of microorganisms of confirmed or potential industrial interest; Biotechnology; Fundamental and applied biological sciences. Psychology; glutaminase; Metabolism. Enzymes; Microbiology; Mission oriented research; Physiology and metabolism; Pseudomonas nitroreducens; γ-glutamyl transferring reaction
• ISSN: 0916-8451; 1347-6947
• DOI: 10.1271/bbb.60.1160

Glutaminase (EC 3.5.1.2) was isolated from Pseudomonas nitroreducens IFO 12694 grown on 0.6% sodium glutamate as a nitrogen source (325-fold purification, 13% yield). The molecular weight of the enzyme was estimated to be 40,000 by gel filtration and SDS-gel electrophoresis. The enzyme hydro-lyzed glutamine optimally at pH 9, and its K m was 6.5 mm. d-Glutamine, γ-glutamyl p-nitroanilide, γ-glutamylmethylamide, γ-glutamylethylamide (theanine), and glutathione showed respectively 107, 85, 78, 74, and 82% reactivity of glutamine. Zn 2+ , Ni 2+ , Cd 2+ , Co 2+ , Fe 2+ , and Cu 2+ repressed the enzyme activity strongly. Glutaminase formed γ-glutamylhydroxamate in the reaction mixture containing glutamine and hydroxylamine (transferring reaction). The optimum pH of the transferring reaction was 7-8, and the K m for glutamine and hydroxylamine were 4 mm and 120 mm, respectively. γ-Glutamyl derivatives hydrolyzable by glutaminase showed reactivity for the transferring reaction. Methylamine or ethylamine was replaceable for hydroxylamine with 3 or 8% reactivity. The effect of divalent cations was not so striking as in the hydrolyzing reaction.