NADP-dependent glutamate dehydrogenase from a facultative methylotroph, Pseudomonas sp. strain AM1.
The NADP-dependent glutamate dehydrogenase (EC 1.4.1.4.) elaborated by the methylotrophic bacterium Pseudomonas sp. strain AM1 when growing on succinate and ammonium chloride was studied. The enzyme, which has a pH optimum of 9.0, was purified 140-fold and shown to have Km values of 20.2 mM, 0.76 mM, 0.033 mM, and 31.6 mM for ammonia, alpha-ketoglutarate, NADPH, and glutamate, respectively. The native molecular weight was determined by polyacrylamide gel electrophoresis to be 190,000, and electrophoresis under denaturing conditions in the presence of sodium dodecyl sulfate revealed a minimum molecular weight of 50,000. The enzyme was highly specific; NADH was unable to replace NADPH in the reaction, various alpha-keto acids could not replace alpha-ketoglutarate, and neither methylamine nor hydroxylamine could substitute for ammonia. Glutamate dehydrogenase was synthesized by the bacteria only when ammonia was its nitrogen source and was repressed if methylamine or nitrate were provided as sources of nitrogen instead of ammonia.
D000643:Ammonium Chloride / Q000378:metabolism; D005969:Glutamate Dehydrogenase / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008744:Methylamines / Q000378:metabolism*; D008970:Molecular Weight; D009249:NADP; D011549:Pseudomonas / Q000201:enzymology*; D013379:Substrate Specificity; D013386:Succinates / Q000378:metabolism; D019802:Succinic Acid
D008744:Methylamines; D013386:Succinates; D000643:Ammonium Chloride; D009249:NADP; D019802:Succinic Acid; C027451:methylamine; D005969:Glutamate Dehydrogenase