title: Unusually stable NAD-specific glutamate dehydrogenase from the alkaliphile Amphibacillus xylanus.
abstract: Nicotinamide adenine dinucleotide-specific glutamate dehydrogenase (NAD-GDH; EC 1.4.1.3) from Amphibacillus xylanus DSM 6626 was enriched 100-fold to homogeneity. The molecular mass was determined by native polyacrylamide electrophoresis and by gel filtration to be 260 kDa (+/- 25 kDa); the enzyme was composed of identical subunits of 45 (+/- 5) kDa, indicating that the native enzyme has a hexameric structure. NAD-GDH was highly specific for the coenzyme NAD(H) and catalyzed both the formation and the oxidation of glutamate. Apparent Km-values of 56 mM glutamate, 0.35 mM NAD (oxidative deamination) and 6.7 mM 2-oxoglutaric acid, 42 mM NH4Cl and 0.036 mM NADH (reductive amination) were measured. The enzyme was unusually resistant towards variation of pH, chaotropic agents, organic solvents, and was stable at elevated temperature, retaining 50% activity after 120 min incubation at 85 degrees C.
authors.affiliation: Fachrichtung 13.3 Mikrobiologie, Universität des Saarlandes Im Stadtwald, Saarbrücken, F.R.G., toja@rz.uni-sb.de
authors.forename: T
authors.initials: T
authors.lastname: Jahns
mesh_terms: D000097:Acetonitriles / Q000494:pharmacology; D000438:Alcohols / Q000494:pharmacology; D000641:Ammonia / Q000378:metabolism; D001407:Bacillus / Q000737:chemistry / Q000201:enzymology*; D001426:Bacterial Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D005969:Glutamate Dehydrogenase / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D018698:Glutamic Acid / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008970:Molecular Weight; D009243:NAD / Q000378:metabolism*; D013696:Temperature
chemical_list: D000097:Acetonitriles; D000438:Alcohols; D001426:Bacterial Proteins; D009243:NAD; D018698:Glutamic Acid; D000641:Ammonia; D005969:Glutamate Dehydrogenase; C032159:acetonitrile