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

---

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

Bovine liver glutamate dehydrogenase reacts covalently with the new adenosine analogue 6-[(4-bromo-2,3-dioxobutyl)thio]-6-deaminoadenosine 5'-diphosphate with incorporation of about 1 mol of reagent/mol of enzyme subunit. Modified enzyme completely loses its normal ability to be inhibited by high concentrations of reduced diphosphopyridine nucleotide (DPNH) (greater than 100 microM), which binds at a regulatory site distinct from the catalytic site; however, the modified enzyme retains its full activity when assayed at 100 microM DPNH in the absence of allosteric compounds. The enzyme is still activated by ADP, is inhibited by GTP (albeit at higher concentrations), and binds 1.5-2 mol of [14C]GTP/subunit. A plot of initial velocity vs. DPNH concentration for the modified enzyme, in contrast to the native enzyme, followed Michaelis-Menten kinetics. The rate constant (k) for loss of DPNH inhibition (as measured at 0.6 mM DPNH) exhibits a nonlinear dependence on reagent concentration, suggesting a reversible binding of reagent (Kd = 0.19 mM) prior to irreversible modification. At 0.1 mM 6-[(4-bromo-2,3-dioxobutyl)thio]-6-deaminoadenosine 5'-diphosphate, k = 0.036 min-1 and is not affected by alpha-ketoglutarate, 100 microM DPNH, or GTP alone but is decreased to 0.0094 min-1 by 5 mM DPNH and essentially to zero by 5 mM DPNH plus 100 microM GTP. Incorporation after incubation with 0.25 mM 6-[(4-bromo-2,3-dioxobutyl)thio]-6-deaminoadenosine 5'-diphosphate for 2 h at pH 7.1 is 1.14 mol/mol of subunit in the absence but only 0.24 mol/mol of subunit in the presence of DPNH plus GTP.(ABSTRACT TRUNCATED AT 250 WORDS)
...
PMID:Affinity labeling of the reduced diphosphopyridine nucleotide inhibitory site of glutamate dehydrogenase by 6-[(4-bromo-2,3-dioxobutyl)thio]-6-deaminoadenosine 5'-diphosphate. 649 69

The digitonin method for the study of cellular compartmentation in mitochondrial and cytosolic fractions was applied to Ehrlich ascites tumor cells. The volume of mitochondrial and cytosolic water spaces are calculated to be 1.62 microliter/30 x 10(6) cells respectively, by the technique of 3H2O permeable and (14C)-sucrose impermeable spaces. The validity of the methods was tested by the distribution of cytosolic (lactate dehydrogenase) and mitochondrial (citrate synthase and glutamate dehydrogenase) marker enzymes. As occurs in normal hepatic cells, an asymmetric distribution of ATP and ADP was observed. The ATP/ADP ratio in the cytosolic fraction was 7 times higher than in the mitochondrial fraction.
...
PMID:Cellular compartmentation of Ehrlich ascites tumor cells. 653 6

Baboons fed ethanol (50% of total calories) chronically develop ultrastructural alterations of hepatic mitochondria. To determine whether mitochondrial functions are also altered, mitochondria were isolated from nine baboons fed ethanol chronically and their pair-fed controls. At the fatty liver stage, ADP-stimulated respiration was depressed in ethanol-fed baboons by 59.4% with glutamate, 43.2% with acetaldehyde, 45.1% with succinate and 51.1% with ascorbate as substrates. A similar decrease was noted in the ADP/O ratio (14 to 28%) and respiratory control ratio (20 to 44%) with all substrates. Similar alterations of mitochondrial functions were observed in baboons with more advanced stages of liver disease, namely fibrosis. These changes after ethanol treatment were associated with decreases in the enzyme activities of mitochondrial respiratory chain: glutamate, NADH and succinate dehydrogenase (42, 24 and 28%, respectively), glutamate-, NADH- or succinate-cytochrome c reductase (42, 27 and 32%, respectively) and cytochrome oxidase (59.6%). The content of all cytochromes was also decreased in ethanol-fed baboons, especially aa3 (57%). Moreover, [14C]leucine incorporation into mitochondrial membranes was depressed by 21% after ethanol treatment. On the other hand, glutamate dehydrogenase activities of serum and cytosol in ethanol-fed baboons were significantly higher than those in pair-fed controls. Morphologically, mitochondria of ethanol-fed baboons were larger than those of pair-fed controls. However, the mitochondrial protein content per mitochondrial DNA was unchanged. From these results, we conclude that, morphologically and functionally, hepatic mitochondria in baboons are altered by chronic ethanol consumption; it is noteworthy that these changes are fully developed already at the fatty liver stage, and that morphological alteration appears to reflect the damage of mitochondrial membranes rather than an adaptive hypertrophy.
...
PMID:Biochemical and morphological alterations of baboon hepatic mitochondria after chronic ethanol consumption. 653 46

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.
...
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.
...
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.
...
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.
...
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.
...
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.
...
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.
...
PMID:The binding of 1,N6-etheno-NAD to bovine liver glutamate dehydrogenase. 674 62


<< Previous 1 2 3 4 5 6 7 8 9 10

---