Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.4.1.2 (glutamate dehydrogenase)
4,380 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Capnocytophaga ochracea strain 25 was originally isolated from a patient with severe juvenile periodontitis. An NAD-specific glutamate dehydrogenase (GDH) (EC 1.4.1.2.) was found in cell-free extracts from this organism. The NADH-dependent reductive, or ammonia-assimilating activity (NADH-GDH), of the enzyme was 8-10-fold higher than its NAD-dependent oxidative, or ammonia-releasing activity (NAD-GDH), suggesting that the primary physiological role of the GDH is ammonia-fixing. Capnocytophaga ochracea GDH was purified approximately 39-fold by a rapid, single-step purification procedure using DEAE-cellulose (DE52) ion-exchange column chromatography which gave 90 per cent recovery of total enzyme units. Paper chromatography of an NADH-GDH assay mixture containing the partially purified enzyme showed that glutamate was, indeed, a product of the ammonia-assimilating reaction. The pH optimum for the NAD-GDH reaction was 9.0; that for the NADH-GDH reaction was 7.5. Although a number of mono- and divalent cations were tested, none had a large effect on either NAD-GDH or NADH-GDH activity. The NAD-GDH reaction showed a hyperbolic kinetic response to glutamate and NAD and the Km values for glutamate and NAD were 2.44 and 0.083 mM respectively. The kinetic response of the NADH-GDH reaction to NADH, alpha-ketoglutarate and ammonium chloride also obeyed Michaelis-Menten kinetics and their respective Km values were 0.069, 1.44 and 3.33 mM. Of a number of biologically-active compounds tested for their ability to modulate GDH activity, only ADP and NAD exerted much effect. The NADH-GDH activity showed a negative hyperbolic kinetic response to both ADP and NAD and Dixon plot-analysis of the NAD and ADP saturation data gave Ki values for ADP and NAD of 4.0 and 0.46 mM respectively. Both NAD and ADP appeared to exert their negative effects on NADH-GDH activity by completely inhibiting the binding of the reduced coenzyme, NADH, to the enzyme.
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PMID:Ammonia utilization by a proposed bacterial pathogen in human periodontal disease, Capnocytophaga ochracea. 657 37

The effect of phosphoenolpyruvate on glutamate dehydrogenase activity was studied in both intact and Triton X-100-treated rabbit renal mitochondria. The intramitochondrial phosphoenolpyruvate content was modulated by application of both 3-MPA, an inhibitor of phosphoenolpyruvate carboxykinase, and BTCA, which inhibits the tricarboxylate-transporting system. The data indicate that: (i) phosphoenolpyruvate is a potent inhibitor of glutamate dehydrogenase activity; and (ii) its inhibitory effect on the enzyme may be abolished by leucine and ADP, activators of glutamate dehydrogenase.
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PMID:Inhibition of glutamate dehydrogenase activity in rabbit renal mitochondria by phosphoenolpyruvate. 662 69

The glutamate dehydrogenase activity found in the serum of patients with Reye's syndrome is shown to be inhibited about 1000-fold more potently by GTP than is the normal human enzyme. 1 mM ADP, which with the normal enzyme effectively reverses GTP inhibition, has no effect in the GTP inhibition of the Reye's syndrome serum activity.
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PMID:Glutamate dehydrogenase in Reye's syndrome. Evidence for the presence of an altered enzyme in serum with increased susceptibility to inhibition by GTP. 663 55

Zone-interference chromatography is a new method for studying macromolecular interactions (S. Endo and A. Wada, Anal. Biochem. 124 (1982) 372). This method is a new style of affinity chromatography which requires no preparation of affinity-column materials but utilizes the velocity difference in a column between interacting molecular species. Using the stochastic theory on the behavior of solute molecules, both the association and the dissociation rate constants can be analytically obtained from the degree of deformation of elution patterns, i.e., the change of the first and second moments. In order to verify the present theory, computer simulation of elution profiles by the extended plate theory and a binding experiment between glutamate dehydrogenase and ADP have been carried out.
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PMID:Theoretical and experimental studies on zone-interference chromatography as a new method for determining macromolecular kinetic constants. 666 97

Glutamic dehydrogenase extracted with tris buffer from fresh freeze-thawed rat heart mitochondria was purified by ammonium sulphate fractionation, affinity chromatography on GTP agarose, hydroxyapatite chromatography and concentration using a molecular sieve. The final specific activity is 80 units/mg protein. Thin gel SDS electrophoresis of the purified enzyme preparation after reduction with dithiothreitol shows a major band with a molecular weight of 38 000 Daltons. Two minor bands are also present. Sucrose density gradient centrifugation reveals a molecular weight of 230 000 Daltons for unreduced mitochondrial GDH activity. By gel filtration rat heart mitochondrial glutamic dehydrogenase has a major peak at 230 000 Daltons, a minor peak at 300 000 Daltons and some larger molecular weight species. Rat liver mitochondrial glutamic dehydrogenase has a minor peak at 230 000, a major peak at 300 000 and some larger molecular weight species. The rat liver mitochondrial glutamic dehydrogenase predominance at 300 000 is unchanged by incubation, extraction and purification with rat heart mitochondria. The purified GDH is stable frozen at -10 degrees C in tris-HCl buffer with EDTA. It loses activity at 4 degrees C especially when stored in 0.2 M phosphate buffer. It also loses activity when dialyzed for 24 h. This loss of activity is not completely prevented by adding nucleotides to the buffer (AMP or ADP) but is decreased by their presence.
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PMID:Glutamic dehydrogenase from rat heart mitochondria. I. Purification and physical properties including molecular weight determination. 672 19

Glutamic dehydrogenase purified from rat heart mitochondria has been characterized with regard to its substrate kinetics and the influence of nucleotides and potassium phosphate on its kinetic properties. The enzyme had characteristics similar to liver mitochondrial glutamic dehydrogenase. These included several double reciprocal plots which were biphasic, indicating homotropic interaction; inhibition by GTP, which was overcome by ADP and phosphate; and activity with both NAD(H) and NADP(H). There were a number of significant differences however, in the specific kinetic properties of heart mitochondrial glutamic dehydrogenase. The Vmax of reductive amination was four-fold greater with NADH than with NADPH. The maximum rate of oxidative deamination was ten-fold greater with NAD compared to NADP. The differences also included: saturating levels of NADH and NADPH were stimulatory rather than inhibitory; ammonia was stimulatory at millimolar levels; NADP and alpha-ketoglutarate were both inhibitory at saturating levels; and ADP increased reductive amination 30% at lower levels of NADH but inhibited at higher (stimulatory) levels of NADH.
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PMID:Glutamic dehydrogenase from rat heart mitochondria. II. Kinetic characteristics. 672 20

The binding of 1,N6-etheno-NAD (epsilon NAD) to bovine liver glutamate dehydrogenase (L-glutamate:NAD(P)+ oxidoreductase (deaminating), EC 1.4.1.3) saturated with glutarate has been investigated at pH 7.0, 0.05 M phosphate buffer at 20 degrees C, by fluorescence titrations. epsilon NAD binds to the protein in a simple fashion: one molecule of coenzyme per enzyme polypeptide chain in the range of enzyme concentrations investigated (from above 50 to a few micromoles of enzyme polypeptide chains/liter). The fluorescence enhancement factor, Q, of bound epsilon NAD relative to free epsilon NAD is independent of the saturation degree, as deduced from the constant value of the long fluorescence decay lifetime (about 21 ns), and is about 17, as deduced from Fmax/F0 ratio values obtained after extrapolation from double reciprocal plots of 1/delta F vs. 1/[glutamate dehydrogenase]. This value for the Q factor is also independent of enzyme concentration, as well as of the presence of either GTP or ADP. At low enzyme concentrations (below 20 mumol polypeptide chains/liter), the dissociation constant of epsilon NAD increases progressively from a plateau value of about 50 microM to about 100 microM at infinite dilution. This is interpreted as being due to a minor affinity of glutamate dehydrogenase hexamers, with respect to higher aggregation states of the enzyme, towards epsilon NAD. As expected, GTP and ADP change the affinity of glutamate dehydrogenase towards epsilon NAD in an opposite manner: GTP strongly increases it, whereas ADP strongly decreases it (Kappd around 6 microM with saturating GTP and around 300 microM with saturating ADP). Furthermore, in the case of GTP, both GTP and epsilon NAD bind to glutamate dehydrogenase with positive cooperativity, with a Hill coefficient of approx. 1.8 for both and a Kappd approximately equal to 30 microM for the binding of GTP to glutamate dehydrogenase saturated with epsilon NAD and glutarate. The value of the Q factor remains the same, even in the presence of the effectors (again from lifetime measurements), as well as the number of epsilon NAD binding sites per enzyme polypeptide chain. These results are interpreted in terms of independent active sites, in the case without effectors. With ADP the binding appears to be simple, but no careful investigation has been attempted at low enzyme concentrations because of the low saturation degree achievable, whereas with GTP the cooperativity can be explained as due to a shift towards hexamers from higher aggregation states.
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PMID:The binding of 1,N6-etheno-NAD to bovine liver glutamate dehydrogenase. 674 62

L-Leucine and its nonmetabolized analogue, 2-aminobicyclo-[2,2,1]heptane-2-carboxylic acid (BCH) activate glutamate dehydrogenase in pancreatic islets, whether the reaction velocity is measured in the direction of glutamate synthesis or glutamate deamination. The rate of glutamate oxidative deamination is increased by ADP and inhibited by 2-ketoglutarate, NH4+ and GTP. The islet homogenate catalyzes the transamination between L-glutamate and either 2-ketoisocaproate or pyruvate, and between 2-ketoglutarate and L-leucine, L-aspartate, L-alanine, L-isoleucine, L-valine, L-norvaline or L-norleucine, but not b (+/-) BCH. The glutamate-aspartate transaminase is preferentially located in mitochondria relative to other transaminases. The parallel effects of L-leucine and BCH on glutamate dehydrogenase and their vastly different abilities to act as transamination partners may account for both analogies and discrepancies in the metabolic and functional responses of the islets to these two branched-chain amino acids.
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PMID:The stimulus-secretion coupling of amino acid-induced insulin release. XI. Kinetics of deamination and transamination reactions. 675 75

1. After chymotryptic digestion of bovine glutamate dehydrogenase, the assay conditions determine whether activation or inhibition is observed. 2. The major fragments appear to remain physically associated. 3. Responses to both GTP and ADP are altered. Inhibition by GTP at pH 7 and 8 is almost abolished. 4. Out of various ligand combinations tested, GTP and NADH together provide the best protection against all the proteolytic effects.
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PMID:Beef liver glutamate dehydrogenase: effects of partial proteolysis with chymotrypsin. 682 6

1. Several glutamate analogues substituted at the beta- or gamma-carbon atoms have been tested as substrates for glutamate dehydrogenase. 2. The two gamma-methyl derivatives and DL-beta-methylglutamate give the same pH optimum (8.7) as L-glutamate, but show inhibition by ADP and activation by GTP as pH 8, unlike glutamate and like the monocarboxylic substrate L-norvaline, which gives a pH optimum of 10. 3. L-gamma-methyleneglutamate, the poorest substrate tested (0.28% of rate with glutamate) gives a high pH optimum (10), like norvaline, but shows marked activation by both ADP (13-fold) and GTP (27-fold). 4. Despite the correct dicarboxylate spacing, all the analogues were much poorer substrates than L-norvaline.
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PMID:Beef liver glutamate dehydrogenase: a study of the oxidation of various alternative amino acid substrates retaining the correct spacing of the two carboxylate groups. 685 48


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