Regulation and properties of glutamine synthetase purified from Bacillus cereus.
The glutamine synthetase from Bacillus cereus IFO 3131 was purified to homogeneity. The enzyme is a dodecamer with a molecular weight of approximately 600,000, and its subunit molecular weight is 50,000. Both Mg2+ and Mn2+ activated the enzyme as to the biosynthesis of L-glutamine, but, unlike in the case of the E. coli enzyme, the Mg2+-dependent activity was stimulated by the addition of Mn2+. The highest activity was obtained when 20 mM Mg2+ and 0.5 mM Mn2+ were added to the assay mixture. For each set of optimal assay conditions, the apparent Km values for glutamate, ammonia and a divalent cation X ATP complex were 1.03, 0.34, and 0.40 mM (Mn2+: ATP = 1: 1); 14.0, 0.47, and 0.91 mM (Mg2+: ATP = 4: 1); and 9.09, 0.45, and 0.77 mM (Mg2+: Mn2+: ATP = 4: 0.2: 1), respectively. At each optimum pH, the Vmax values for these reactions were 6.1 (Mn2+-dependent), 7.4 (Mg2+-dependent), and 12.9 (Mg2+ plus Mn2+-dependent) mumoles per min per mg protein, respectively. Mg2+-dependent glutamine synthetase activity was inhibited by the addition of AMP or glutamine; however, this inhibitory effect was suppressed in the case of the Mg2+ plus Mn2+-dependent reaction. These results suggest that the activity of the B. cereus glutamine synthetase is regulated by both the intracellular concentration and the ratio of Mn2+/Mg2+ in vivo. Also in the present investigation, a potent glutamine synthetase inhibitor(s) was detected in crude extracts from B. cereus.
Matsuoka
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Kurebayashi
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Kimura
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D000645:Ammonium Sulfate; D001409:Bacillus cereus / Q000201:enzymology*; D002384:Catalysis; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation / Q000187:drug effects; D005974:Glutamate-Ammonia Ligase / Q000302:isolation & purification / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D006882:Hydroxyapatites; D066298:In Vitro Techniques; D008970:Molecular Weight; D013307:Streptomycin / Q000494:pharmacology
D006882:Hydroxyapatites; D005974:Glutamate-Ammonia Ligase; D000645:Ammonium Sulfate; D013307:Streptomycin