Purification and properties of glutamate synthase and glutamate dehydrogenase from Bacillus megaterium.
نویسندگان
چکیده
Bacillus megaterium N.C.T.C. no. 10342 exhibits glutamate synthetase (EC 2.6.1.53) and glutamate dehydrogenase (EC 1.4.1.4) activities. Concentrations of glutamate synthase were high when the bacteria were grown on 3mM-NH4Cl and low when they were grown on 100mM-NH4Cl, whereas glutamate dehydrogenase concentrations were higher when the bacteria were grown on 100mM-NH4Cl than on 3mM-NH4Cl. Glutamate synthase and glutamate dehydrogenase were purified to homogeneity from B. megaterium grown in 10mM-glucose/10mM-NH4Cl. The purified enzymes had mol.wts. 840000 and 270000 for glutamate synthase and glutamate dehydrogenase respectively. The Km values for substrates with NADPH and coenzyme were (glutamate synthase activity shown first) 9 micron and 360 micron for 2-oxoglutarate, 7.1 micron and 8.7 micron for NADPH, and 0.2 mM for glutamine and 22 mM for NH4Cl, similar values to those of enzymes from Escherichia coli. Glutamate synthase contained NH3-dependent activity (different from authentic glutamate dehydrogenase), which was enhanced 4-fold during treatment at pH 4.6 NH3-dependent activity was generally about 2% of the glutamine-dependent activity. Amidination of glutamate synthase by the bi-functional cross-linking reagent dimethyl suberimidate inactivated glutamine-dependent glutamate synthase activity, but increased NH3-dependent activity. A cross-linked structure of mol.wt. approx 200000 was the main product formed.
منابع مشابه
Expression Optimization and Purification of Glutamate Endopeptidase from halo-thermo tolerant Bacillus licheniformis SL-1
Aims and Background: Glutamate-specific endopeptidase (GSE, EC 3.4.21.19) is belonging to the serine protease family enzymes. A glutamate-specific endopeptidase has the ability to cleavage peptide bonds for the protein structure analysis, solid phase synthesis of peptides and preparation of nanotubes. The purpose of this investigation was to produce glutamate endopeptidase enzyme from the Bacil...
متن کامل13N isotope studies on the pathway of ammonia assimilation in Bacillus megaterium and Escherichia coli.
The pathway of ammonia incorporation into amino acids was studied by use of 13N-ammonium ions in Bacillus megaterium and Escherichia coli that had been grown aerobically on a minimal salts medium containing NH4Cl as the source of nitrogen. Anion- and cation-exchange high-pressure-liquid chromatography was used to separate amino acids relevant to the several possible pathways for ammonia assimil...
متن کاملGlutamate biosynthesis in Bacillus azotofixans. 15N NMR and enzymatic studies.
Pathways of ammonia assimilation into glutamic acid in Bacillus azotofixans, a recently characterized nitrogen-fixing species of Bacillus, were investigated through observation by NMR spectroscopy of in vivo incorporation of 15N into glutamine and glutamic acid in the absence and presence of inhibitors of ammonia-assimilating enzymes, in combination with measurements of the specific activities ...
متن کاملPurification and Chemical Characterization of Malate Dehydrogenase of Bacillus subtilis
In an attempt to elucidate the relationships between some pyridine nucleotide-dependent dehydrogenases in Bacillus subtilis, such as alanine dehydrogenase, malate dehydrogenase, lactate dehydrogenase, and glutamate dehydrogenase, these enzymes are being purified and characterized with respect to their chemical, serological, and enzymic properties. In two previous papers (1, 2), the purification...
متن کاملHepatic Glutamate Dehydrogenase Activity and the Presence of Ammonia and Urea in the Circulatory Fluid of Channa Gachua in Water-Restricted Condition
Ammonia is the chief excretory product in fishes. However, non-availability of enough of water in the habitat, may lead to the formation of urea, in fishes. In the present study, the possible role of urea formation to avoid the toxicity of ammonia under water-restricted condition was tested in Channa gachua. Circulatory urea and ammonia were estimated in the blood of the fishes and glu...
متن کاملذخیره در منابع من
با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید
برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید
ثبت ناماگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید
ورودعنوان ژورنال:
- The Biochemical journal
دوره 173 1 شماره
صفحات -
تاریخ انتشار 1978