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)

The reaction mechanism of Azospirillum brasilense glutamate synthase has been investigated by several approaches. 15N nuclear magnetic resonance studies demonstrate that the amide nitrogen of glutamine is reductively transferred to 2-oxoglutarate in an irreversible manner with no release of the transferred ammonia group into the medium. Identical results were obtained using thio-NADPH and acetylpyridine-NADPH, which are shown to be less efficient substrates of the enzyme than NADPH. Similarly, no exchange of the ammonia group being transferred with exogenous ammonium ion was observed during catalysis. The glutamate formed as the product of the iminoglutarate reduction was determined to be in the L configuration. The enzyme was also found to catalyze, under anaerobic conditions, the exchange of the 4proS H of NADPH with solvent both in the absence and in the presence of 2-oxoglutarate and glutamine. The reductive half-reaction is therefore a reversible segment of the overall irreversible amidotransferase reaction. 15N NMR studies also showed that the enzyme does not catalyze glutamate dehydrogenase/oxidase reactions or any observable glutaminase activity under neutral (pH 7.5) conditions. Glutaminase activity was also not observable with the reduced enzyme alone or in the presence of D-glutamate (a competitive inhibitor of glutamate synthase with respect to 2-oxoglutarate, with a Ki of about 11 microM) or with the oxidized enzyme in the presence of 2-oxoglutarate, D-glutamate, or NADP+. These data confirm species-dependent differences of A. brasilense glutamate synthase with respect to the enzyme from other sources.
Biochemistry 1991 Dec 03
PMID:Mechanistic studies on Azospirillum brasilense glutamate synthase. 168 91

A 52 kDa protein could only be co-purified with the CoA-modified forms of acetyl-CoA acetyltransferase (acetoacetyl-CoA thiolase) (EC 2.3.1.9) from rat liver mitochondria. Immunoprecipitations of these modified forms with anti-(acetyl-CoA acetyltransferase) IgG or anti-(52 kDa protein) IgG yielded, in addition to the appropriate proteins, the 52 kDa protein or the CoA-modified form of acetyl-CoA acetyltransferase (41 kDa) respectively. This was demonstrated by SDS/PAGE and immunoblots. The modified forms containing the 52 kDa protein could be cross-linked by 1,5-difluoro-2,4-dinitrobenzene to a high-molecular-mass complex containing both the 41 kDa and 52 kDa proteins. The 52 kDa protein was identified as mitochondrial glutamate dehydrogenase (EC 1.4.1.3) by amino acid sequence analysis. The results of co-immunoprecipitation and cross-linking characterize the CoA-modified forms of acetyl-CoA acetyltransferase and the glutamate dehydrogenase as nearest-neighbour proteins.
Biochem J 1991 Dec 01
PMID:Identification of the CoA-modified forms of mitochondrial acetyl-CoA acetyltransferase and of glutamate dehydrogenase as nearest-neighbour proteins. 168 1

Progressive changes in serum enzyme activity and liver histologic features were monitored in calves fed tansy ragwort (Senecio jacobaea)-contaminated pellets. The experiments were designed to simulate natural intoxicant ingestion conditions in relationship to the dose and duration of exposure to the toxic plant to correlate early laboratory diagnostic changes with the natural progression of the disease, thereby facilitating early diagnosis and intervention by veterinary clinicians. Eight calves were fed tansy ragwort and 4 additional calves served as controls. In group 1, 4 calves were continuously fed dried tansy ragwort mixed in a pelleted feed at a 5% concentration by dry weight until terminal liver disease developed. Serum liver enzyme (alkaline phosphatase, glutamate dehydrogenase, and gamma-glutamyltransferase) activities were monitored at weekly intervals in these calves and in the 2 controls. In group 2, 4 calves were fed the same contaminated feed for only 60 days, with return to normal feed for the duration of the trial. Two additional calves served as controls. Their liver enzyme activities were monitored every other week in conjunction with percutaneous liver biopsies. All 8 calves fed tansy ragwort-contaminated pellets developed terminal hepatopathy in either a chronic pattern (n = 6) or a chronic-delayed pattern (n = 2), with the onset of a moribund state or sudden death at 11 to 17 weeks and 27 to 51 weeks, respectively. The calves were euthanatized when classic terminal signs of hepatic encephalopathy first became evident. The clinicopathologic patterns of chronic and chronic-delayed toxicoses were typical of over 5,000 cases of field tansy toxicosis diagnosed at the diagnostic laboratory. Serum glutamate dehydrogenase was the first enzyme to increase in most animals, with a short-term increase to peak values followed by a rapid return to normal. This enzyme change was followed by increases in alkaline phosphatase and gamma-glutamyltransferase. Serum enzyme changes preceded development of recognizable histologic lesions. Vacuolar changes in hepatocyte nuclei, biliary hyperplasia, and fibrosis sequentially developed in liver biopsy specimens from each animal, whereas megalocytosis was not a predominant feature until necropsy. On the basis of our findings, we suggest that the optimal tests for diagnosis of pyrrolizidine alkaloid intoxication should consist of liver biopsy and determination of concurrent serum liver-enzyme activities.
Am J Vet Res 1991 Dec
PMID:Serum liver enzyme and histopathologic changes in calves with chronic and chronic-delayed Senecio jacobaea toxicosis. 168 78

Treatment of rats with the vitamin B12 analogue hydroxy-cobalamin[c-lactam] (HCCL) impairs methylmalonyl-CoA mutase function and leads to methylmalonic aciduria due to intracellular accumulation of propionyl and methylmalonyl-CoA. Since accumulation of these acyl-CoAs disrupts normal cellular regulation, the present investigation characterized metabolism in hepatocytes and liver mitochondria from rats treated subcutaneously with HCCL or saline (control) by osmotic minipump. Consistent with decreased methylmalonyl-CoA mutase activity, 14CO2 production from 1-14C-propionate (1 mM) was decreased by 76% and 82% after 2-3 wk and 5-6 wk of HCCL treatment, respectively. In contrast, after 5-6 wk of HCCL treatment, 14CO2 production from 1-14C-pyruvate (10 mM) and 1-14C-palmitate (0.8 mM) were increased by 45% and 49%, respectively. In isolated liver mitochondria, state 3 oxidation rates were unchanged or decreased, and activities of the mitochondrial enzymes, citrate synthetase, succinate dehydrogenase, carnitine palmitoyltransferase, and glutamate dehydrogenase (expressed per milligram mitochondrial protein) were unaffected by HCCL treatment. In contrast, activities of the same enzymes were significantly increased in both liver homogenate (expressed per gram liver) and isolated hepatocytes (expressed per 10(6) cells) from HCCL-treated rats. The mitochondrial protein per gram liver, calculated on the basis of the recovery of the mitochondrial enzymes, increased by 39% in 5-6 wk HCCL-treated rats. Activities of lactate dehydrogenase, catalase, cyanide-insensitive palmitoyl-CoA oxidation, and arylsulfatase A in liver were not affected by HCCL treatment. Hepatic levels of mitochondrial mRNAs were elevated up to 10-fold in HCCL-treated animals as assessed by Northern blot analysis. Thus, HCCL treatment is associated with enhanced mitochondrial oxidative capacity and an increased mitochondrial protein content per gram liver. Increased mitochondrial oxidative capacity may be a compensatory mechanism in response to the metabolic insult induced by HCCL administration.
J Clin Invest 1990 Dec
PMID:Increased hepatic mitochondrial capacity in rats with hydroxy-cobalamin[c-lactam]-induced methylmalonic aciduria. 170 51

The neuropathological findings in a patient with antemortem diagnosis of olivopontocerebellar atrophy (OPCA) and reduced leucocytic glutamate dehydrogenase (GDH) activity included cerebellar cortical degeneration, most marked in the superior vermis, mild atrophy of the pons and the inferior olivary nucleus, marked reduction of anterior horn cells at all levels and gliosis in both lateral columns. GDH activities and their thermolability in "soluble" and "particulate" fractions in the cerebral cortex, cerebellar hemisphere and vermis were not significantly different from the values in two control brains. GDH mRNA in the patient's brain was not altered in size or amount.
J Neurol Neurosurg Psychiatry 1990 Dec
PMID:Multiple system degeneration with glutamate dehydrogenase deficiency: pathology and biochemistry. 170 75

We propose a testable general mechanism by which ligand binding energy can be used to drive a catalytic step in an enzyme catalyzed reaction or to do other forms of work involving protein molecules. This energy transduction theory is based on our finding of the widespread occurrence of ligand binding-induced protein macrostate interconversions each having a large invariant delta H0 accompanied by a small but highly variable delta G0. This phenomenon, which can be recognized by the large delta Cp0's it generates, can provide the necessary energy input step but is not in itself sufficient to constitute a workable transduction mechanism. A viable mechanism requires the additional presence of an 'energy transmission step' which is terminated to trigger the 'power' stroke at a precise location on the reaction coordinate, followed by an energetically inexpensive 'return' step to restore the machine to its initial conditions. In the model we propose here, these additional steps are provided by the existence of ligand inducible 2-state transitions in the free enzyme and in each of the enzyme complexes that occur along the reaction coordinate, and by the selective blocking of certain of these interconversions by high energetic barriers. We provide direct experimental evidence supporting the facts that these additional mechanistic components do exist and that the liver glutamate dehydrogenase reaction is indeed driven by just such machinery. We describe some aspects of the chemical nature of these transitions, and evidence for their occurrence in other systems.
FEBS Lett 1991 Dec 02
PMID:Transduction of enzyme-ligand binding energy into catalytic driving force. 174 79

Steady-state kinetic properties of glutamate dehydrogenase from Clostridium symbiosum are reported. Rates with NADP(H) are over three hundred times lower than with NAD(H) under identical conditions. The 3-acetyl pyridine and 6-deamino adenine analogues of NAD+, on the other hand, are used almost as well as NAD+ itself. Amino acid specificity is very tight at both pH 7 and pH 9. The best alternative substrate of those tested, L-alpha-amino-gamma-nitraminobutyrate, gave only 0.5% of the rate seen with glutamate. With 400 microM NAD+ a 160-fold variation of the glutamate concentration gave a linear Eadie plot apart from slight inhibition at the highest concentrations. With 40 mM L-glutamate and varied [NAD+], the Eadie plot appeared linear between 1.6 microM and 60 microM and again between 60 microM and 2000 microM, but the slopes of the two lines differed by a factor of 8.4. This striking pattern is not attributable to impurities in the coenzyme or to changes in the state of aggregation of the enzyme. For the high concentration range (greater than 60 microM NAD+), the presence of all four linear terms in the reciprocal form of the initial rate equation indicates a sequential mechanism. Similar measurements made for APAD+ and dnNAD+ show no sign of non-linearity in the Eadie plot over the wide concentration ranges explored. In the reductive amination direction, with NADH as coenzyme, linear reciprocal plots were obtained for all three substrates. Systematic variation of concentrations led via primary, secondary and tertiary plots to all eight possible initial-rate parameters in a linear reciprocal initial-rate equation. Compulsory-order and enzyme-substitution mechanisms appear to be excluded, and a random route to the central complex seems the only possibility compatible with the results.
Biochim Biophys Acta 1991 Dec 06
PMID:Functional studies of a glutamate dehydrogenase with known three-dimensional structure: steady-state kinetics of the forward and reverse reactions catalysed by the NAD(+)-dependent glutamate dehydrogenase of Clostridium symbiosum. 176 63

The hyperthermophilic archaebacterium Pyrococcus furiosus contains high levels of NAD(P)-dependent glutamate dehydrogenase activity. The enzyme could be involved in the first step of nitrogen metabolism, catalyzing the conversion of 2-oxoglutarate and ammonia to glutamate. The enzyme, purified to homogeneity, is a hexamer of 290 kDa (subunit mass 48 kDa). Isoelectric-focusing analysis of the purified enzyme showed a pI of 4.5. The enzyme shows strict specificity for 2-oxoglutarate and L-glutamate but utilizes both NADH and NADPH as cofactors. The purified enzyme reveals an outstanding thermal stability (the half-life for thermal inactivation at 100 degrees C was 12 h), totally independent of enzyme concentration. P. furiosus glutamate dehydrogenase represents 20% of the total protein; this elevated concentration raises questions about the roles of this enzyme in the metabolism of P. furiosus.
Eur J Biochem 1991 Dec 18
PMID:Extremely thermostable glutamate dehydrogenase from the hyperthermophilic archaebacterium Pyrococcus furiosus. 176 79

We have isolated and sequenced the gene for a putative NADP-dependent glutamate dehydrogenase from the extremely halophilic archaebacterium Halobacterium salinarium. This gene is transcribed as a unique RNA molecule of about 1700 nucleotides. The 5' end of the transcript contains characteristic consensus transcription initiation and promoter sequences observed in halophilic archaebacteria. The encoded polypeptide, with a predicted length of 435 amino acids, shows significant overall homology and conservation of functional domains when compared with different eubacterial and eukaryotic glutamate dehydrogenases. Surprisingly, the archaebacterial protein shares a larger number of identical amino acid residues with homologous polypeptides from higher eukaryotes than with those from unicellular eukaryotes and eubacteria.
Mol Gen Genet 1991 Dec
PMID:The gene for a halophilic glutamate dehydrogenase: sequence, transcription analysis and phylogenetic implications. 176 32

Pregnant rats of 19th and 21st days were given an acute nitrogen overload produced by an infusion of either 0.2 M ammonium acetate or 0.2 M glutamine. Metabolic adaptations to nitrogen excess were studied measuring--in fetomaternal unit--non-protein nitrogen content and the activities of enzymes related with ammonia metabolism. Maternal and fetal plasma urea levels were increased by ammonium acetate treatment. Glutamine overload increased more the amino acid content in the mothers than in conceptus. As response to ammonium acetate treatment, glutamate dehydrogenase activity in liver was more sensitive in pregnant than in nonpregnant rats, suggesting more nitrogen incorporation into amino acids in pregnancy. Regarding glutamine synthetase activity, both treatments had an opposite effect except in kidney. The adenylate deaminase activity of pregnant rats was inhibited similarly to nonpregnant rats by nitrogen overloads, but stronger after glutamine infusion. Placenta and fetal metabolism were adjusted, as the dams, to lack of ammonia production by nitrogen overloads and to glutamine synthesis by ammonium acetate infusion.
Horm Metab Res 1991 Dec
PMID:Metabolic adaptations to nitrogen excess in late gestation in rat. 177 94


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