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)

We have analysed the sequence homology between glutamate, leucine and phenylalanine dehydrogenases in the light of the solution of the structure of the glutamate dehydrogenase from Clostridium symbiosum. This analysis indicates that the elements of secondary structure comprising the core of the two domains in glutamate dehydrogenase are conserved in the other two enzymes. There is a striking conservation of the residues responsible for the recognition of the nicotinamide ring of the nucleotide cofactor and the backbone of the amino acid substrates. Furthermore, residues involved in a major conformational rearrangement on amino acid binding are preserved, as are those implicated in the catalytic chemistry. In contrast, the pattern of insertions/deletions between these enzymes is consistent with possible differences in quaternary structure. Differential substrate specificity between these enzymes is achieved by critical substitutions at the base of the binding pocket, which accommodates the side-chain of the amino acid substrate. This provides insights into the mutations necessary to produce new catalysts for the chiral synthesis of novel amino acids.
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PMID:Evolution of substrate diversity in the superfamily of amino acid dehydrogenases. Prospects for rational chiral synthesis. 826 39

Although heart mitochondria contain glutamate dehydrogenase, it has not been thought to play a role in their metabolism. We investigated this matter to define the conditions under which it is active. We found modest activity in the presence of glutamate and malate and a continuous source of ADP when pyruvate is added. This increases several fold as the osmolarity is increased from 296 to 370 mosM. At the higher osmolarity ammonia formation is brief, associated with a lower intramitochondrial alpha-ketoglutarate from citrate does not make up for the drop in glutamate conversion to alpha-ketoglutarate. Mitochondrial content of nucleotides and CoA compounds are not altered by pyruvate addition. The rate of glutamate deamination by GDH in sonicated heart mitochondria agrees with the rate of ammonia formation in intact mitochondria in the presence of pyruvate (20 nmol/min/mg of mitochondrial protein). We conclude pyruvate lowers mitochondrial oxalacetate which decreases alpha-ketoglutarate formation by transamination. The lower mitochondria alpha-ketoglutarate level permits glutamate deamination until alpha-ketoglutarate reaches a level that inhibits the forward reaction. Further proof of the key role of alpha-ketoglutarate is seen with aminooxyacetate which blocks transamination. In its presence ammonia formation occurs at the same rate (18 nm/min/mg of mitochondrial protein), is not dependent upon pyruvate, and does not stop after a couple of minutes. Leucine, which decreases alpha-ketoglutarate inhibition of GDH, also results in ammonia formation, further supporting the concept of regulation by alpha-ketoglutarate. The higher osmolarity increases GDH activity by increasing alpha-ketoglutarate transport from mitochondria.
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PMID:Conditions for glutamate dehydrogenase activity in heart mitochondria. 837 37

The carboxyl-terminal catalytic domain of the human poly(ADP-ribose) polymerase (PARP) exhibits sequence homology with the NAD(P)(+)-dependent leucine and glutamate dehydrogenases. To clarify the role played by some conserved residues between PARP and NAD(P)(+)-dependent dehydrogenases, point mutations were introduced into the whole enzyme context. Non-conservative mutations of Lys-893 (K893I) and Asp-993 (D993A) completely inactivate human PARP, whereas conservative and nonconservative mutations of Asp-914 (D914E and D914A, respectively) and Lys-953 (K953R and K953I, respectively) partially alter PARP activity. The consequences of conservative substitution of Lys-893 and Asp-993 on the kinetic properties of human poly(ADP-ribose) polymerase enzyme and the polymer it synthesizes suggest that these 2 amino acids are directly involved in the covalent attachment of the first ADP-ribosyl residue from NAD+ onto the acceptor amino acid. In addition, the recent resolution of the three-dimensional structure of the NAD(+)-linked glutamate dehydrogenase from Clostridium symbiosum (Baker, P.J., Britton, K.L., Engel, P.C., Farrants, G.W., Lilley, K.S., Rice, D.W., and Stillman, T.J. (1992) Proteins 12, 75-86) strongly supports our alignment with leucine and glutamate dehydrogenases and provides an interesting structural framework for the analysis of our results of site-directed mutagenesis.
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PMID:Identification of potential active-site residues in the human poly(ADP-ribose) polymerase. 847 97

Gabapentin is a novel anticonvulsant drug. The anticonvulsant mechanism of gabapentin is not known. Based on the amino acid structure of gabapentin we explored its possible effects on glutamate and gamma-aminobutyric acid (GABA) metabolism in brain as they may relate to its anticonvulsant mechanisms of action. Gabapentin was tested for its effects on seven enzymes in the metabolic pathways of these two neurotransmitters: alanine aminotransferase (AL-T), aspartate aminotransferase (AS-T), GABA aminotransferase (GABA-T), branched-chain amino acid aminotransferase (BCAA-T), glutamine synthetase (Gln-S), glutaminase (GLNase), and glutamate dehydrogenase (GDH). In the presence of 10 mM gabapentin, only GABA-T, BCAA-T, and GDH activities were affected by this drug. Inhibition of GABA-T by gabapentin was weak (33%). The Ki values for inhibition of cytosolic and mitochondrial forms of GABA-T (17-20 mM) were much higher than the Km values for GABA (1.5-1.9 mM). It is, therefore, unlikely that inhibition of GABA-T by gabapentin is clinically relevant. As with leucine, gabapentin stimulated GDH activity. The GDH activity in rat brain synaptosomes was activated 6-fold and 3.4-fold, respectively, at saturating concentrations (10 mM) of leucine and gabapentin. The half-maximal stimulation by gabapentin was observed at approximately 1.5 mM. Gabapentin is not a substrate of BCAA-T, but it exhibited a potent competitive inhibition of both cytosolic and mitochondrial forms of brain BCAA-T. Inhibition of BCAA-T by this drug was reversible. The Ki values (0.8-1.4 mM) for inhibition of transamination by gabapentin were close to the apparent Km values for the branched-chain amino acids (BCAA) L-leucine, L-isoleucine, and L-valine (0.6-1.2 mM), suggesting that gabapentin may significantly reduce synthesis of glutamate from BCAA in brain by acting on BCAA-T.
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PMID:Effects of anticonvulsant drug gabapentin on the enzymes in metabolic pathways of glutamate and GABA. 856 62

To date, no attempt has been made to study alterations occurring in the amino acid profile in chronic models of thioacetamide-induced liver cirrhosis. In this work, changes in serum amino acids and proteins in rats with thioacetamide-induced liver cirrhosis are reported, together with changes in enzyme activities in the liver and serum. Seventeen female Wistar rats were used. Eight rats were given 300 mg thioacetamide/l in drinking water for 4 months and nine rats were given water ad libitum during the same time-period. Significant increases in glycine, alanine, serine, methionine, glutamate, ornithine, phenylalanine, tyrosine, histidine and proline were observed in rats with the resulting experimental liver cirrhosis. Threonine, taurine, glutamine, lysine and citrulline tended to increase while isoleucine, leucine, aspartate, arginine and tryptophan tended to decrease. Total and nonessential amino acids increased significantly in cirrhotic animals. Total essential and aromatic amino acids tended to increase in the thioacetamide-treated group, whereas branched chain amino acids tended to decrease in the same group. Regarding serum proteins, a decrease in albumin concentration in the thioacetamide-treated animals was the only change detected. The liver enzyme activities under observation (aspartate and alanine aminotransferases, glutamate dehydrogenase and threonine deaminase) were lower in the thioacetamide group. Decreases were significant for both transaminases and threonine deaminase. Results for serum activities showed that transaminases did not change in thioacetamide-treated rats in comparison with controls. In contrast, alkaline phosphatase rose dramatically in cirrhotic rats. We conclude that the serum amino acid pattern in this chronic model of liver cirrhosis resembles in part that of the corresponding human disease.
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PMID:Serum amino acid changes in rats with thioacetamide-induced liver cirrhosis. 857 92

Little information is available regarding the regulation of glutamate dehydrogenase in skeletal muscle. We investigated the regulation of glutamate dehydrogenase by branched-chain amino acids (BCAA) in skeletal muscles from rats and chicks and determined the effects of metabolic acidosis on the activity and regulation of this enzyme by BCAA in rat skeletal muscle. Skeletal muscle mitochondria were prepared from normal rats and chicks and acidotic rats. Mitochondrial glutamate dehydrogenase activity was measured in the presence or absence of BCAA. Metabolic acidosis was induced by feeding rats 1.5% NH4Cl as drinking water. Glutamate dehydrogenase activity was stimulated by leucine (P < 0.001) and isoleucine (P < 0.05) in rat muscles and by leucine (P < 0.05) in chick muscles in a concentration-dependent manner. Both leucine and isoleucine had their maximum effects at a concentration of 1 mM (45% by leucine and 27% by isoleucine in rat muscle; 36% by leucine in chick muscle). The maximum stimulatory effects of leucine and isoleucine in rat muscles were additive. Neither valine nor 2-oxoisocaproate had an effect on glutamate dehydrogenase activity in rat or chick muscles. In acidotic rats, the basal activity of skeletal muscle glutamate dehydrogenase was 1.8-fold (P < 0.01) greater than in control rats; leucine, isoleucine, and valine significantly increased glutamate dehydrogenase activity (maximally 86, 55 and 33%, respectively; P < 0.05). We conclude that glutamate dehydrogenase activity in skeletal muscle from rats and chicks is regulated by BCAA, and that a species difference exists between rats and chicks. Metabolic acidosis increases the activity of glutamate dehydrogenase and its sensitivity to BCAA.
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PMID:Regulation of glutamate dehydrogenase by branched-chain amino acids in skeletal muscle from rats and chicks. 892 9

We tested the hypothesis that nutritional state affects seawater acclimation by transferring either fed or food-deprived (2 weeks) male tilapia (Oreochromis mossambicus) from fresh water to full-strength sea water. Food-deprivation resulted in a significant increase in plasma concentrations of Na+, Cl-, cortisol, glucose, total amino acid, glutamate, serine and alanine, and in hepatic pyruvate kinase (PK) and lactate dehydrogenase (LDH) activities, whereas the prolactin-188 to prolactin-177 ratio (tPRL188:tPRL177) and plasma prolactin-188 (tPRL188), lactate, arginine and hepatic glycogen content and hepatic alanine aminotransferase (AlaAT) and 3-hydroxyacyl-Coenzyme A dehydrogenase (HOAD) activities were lower than in the fed group. Seawater transfer significantly increased the tPRL188:tPRL177 ratio and plasma concentrations of Na+, Cl-, K+, growth hormone (GH), glucose, aspartate, tyrosine, alanine, methionine, phenylalanine, leucine, isoleucine and valine levels as well as gill Na+/K+-ATPase activity and hepatic PK and LDH activities, whereas plasma tPRL177, tPRL188, glycine and lysine concentrations were significantly lower than in fish retained in fresh water. There was a significant interaction between nutritional state and salinity that affected the tPRL188:tPRL177 ratio and plasma concentrations of Cl-, GH, glucose, aspartate, tyrosine, serine, alanine, glycine, arginine and hepatic PK, LDH, AlaAT, aspartate aminotransferase, glutamate dehydrogenase and HOAD activities. These results, taken together, indicate that food-deprived fish did not regulate their plasma Cl- levels, despite an enhancement of plasma hormonal and metabolic responses in sea water. Our study also suggests the possibility that plasma prolactin and essential amino acids may be playing an important role in the seawater acclimation process in tilapia.
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PMID:Food-deprivation affects seawater acclimation in tilapia: hormonal and metabolic changes 932 Mar 94

The subunit of the enzyme glutamate dehydrogenase comprises two domains separated by a cleft harboring the active site. One domain is responsible for dinucleotide binding and the other carries the majority of residues which bind the substrate. During the catalytic cycle a large movement between the two domains occurs, closing the cleft and bringing the C4 of the nicotinamide ring and the Calpha of the substrate into the correct positioning for hydride transfer. In the active site, two residues, K89 and S380, make interactions with the gamma-carboxyl group of the glutamate substrate. In leucine dehydrogenase, an enzyme belonging to the same superfamily, the equivalent residues are L40 and V294, which create a more hydrophobic specificity pocket and provide an explanation for their differential substrate specificity. In an attempt to change the substrate specificity of glutamate dehydrogenase toward that of leucine dehydrogenase, a double mutant, K89L,S380V, of glutamate dehydrogenase has been constructed. Far from having a high specificity for leucine, this mutant appears to be devoid of any catalytic activity over a wide range of substrates tested. Determination of the three-dimensional structure of the mutant enzyme has shown that the loss of function is related to a disordering of residues linking the enzyme's two domains, probably arising from a steric clash between the valine side chain, introduced at position 380 in the mutant, and a conserved threonine residue, T193. In leucine dehydrogenase the steric clash between the equivalent valine and threonine side chains (V294, T134) does not occur owing to shifts of the main chain to which these side chains are attached. Thus, the differential substrate specificity seen in the amino acid dehydrogenase superfamily arises from both the introduction of simple point mutations and the fine tuning of the active site pocket defined by small but significant main chain rearrangements.
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PMID:Determinants of substrate specificity in the superfamily of amino acid dehydrogenases. 940 44

Klebsiella aerogenes strains with reduced levels of D-amino acid dehydrogenase not only fail to use alanine as a growth substrate but also become sensitive to alanine in minimal media supplemented with glucose and ammonium. The inability of these mutant strains to catabolize the alanine provided in the medium interferes with both pathways of glutamate production. Alanine derepresses the nitrogen regulatory system (Ntr), which in turn represses glutamate dehydrogenase, one pathway of glutamate production. Alanine also inhibits the enzyme glutamine synthetase, the first enzyme in the other pathway of glutamate production. Therefore, in the presence of alanine, strains with mutations in dadA (the gene that codes for a subunit of the dehydrogenase) exhibit a glutamate auxotrophy when ammonium is the sole source of nitrogen. The alanine catabolic operon of Klebsiella aerogenes, dadAB, was cloned, and its DNA sequence was determined. The clone complemented the alanine defects of dadA strains. The operon has a high similarity to the dadAB operon of Salmonella typhimurium and the dadAX operon of Escherichia coli, each of which codes for the smaller subunit of D-amino acid dehydrogenase and the catabolic alanine racemase. Unlike the cases for E. coli and S. typhimurium, the dad operon of K. aerogenes is activated by the Ntr system, mediated in this case by the nitrogen assimilation control protein (NAC). A sequence matching the DNA consensus for NAC-binding sites is located centered at position -44 with respect to the start of transcription. The promoter of this operon also contains consensus binding sites for the catabolite activator protein and the leucine-responsive regulatory protein.
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PMID:Alanine catabolism in Klebsiella aerogenes: molecular characterization of the dadAB operon and its regulation by the nitrogen assimilation control protein. 945 58

The ability of alpha-ketoisocaproate (KIC) to induce ATP production in isolated mitochondria from pancreatic beta-cells was examined with a bioluminometric method. There was no ATP production from KIC when tested alone or in combination with malate (1 mmol/l), nor did DL-beta-hydroxybutyrate induce mitochondrial ATP production, whereas palmitoyl-carnitine and pyruvate were efficient stimulators of mitochondrial ATP production in the presence of an equimolar concentration of malate. However, KIC stimulated the mitochondrial ATP production when tested in combination with glutamate (10 mmol/l). The concentration necessary to obtain half-maximal stimulation was approximately 50 micromol/l KIC, and maximal activity, comparable to that obtained with fatty acids, was reached at 1 mmol/l KIC. Higher KIC concentrations inhibited the mitochondrial ATP production, whereas a plateau was attained at 1 mmol/l KIC in the presence of glutamine. Ca2+ stimulated the maximal mitochondrial ATP production induced by KIC. Maximal stimulation was obtained with 300 nmol/l Ca2+ in the presence of 0.3 mmol/l KIC. Ca2+ reduced the concentration of KIC necessary for half-maximal stimulation to <30 micromol/l. Leucine stimulated the mitochondrial ATP production in the presence of glutamate to the same extent as KIC. Half-maximal stimulation was observed with 2 mmol/l leucine. There were no additive effects on mitochondrial ATP production when KIC and leucine were tested in combination. The results demonstrate that KIC by itself is not a mitochondrial substrate for ATP production. KIC must transaminate with glutamate or glutamine to yield alpha-ketoglutarate and leucine. Since leucine allosterically activates glutamate dehydrogenase, which also produces alpha-ketoglutarate, the insulinogenic effect of KIC may in part be due to the intramitochondrial generation of alpha-ketoglutarate. Since KIC-induced ATP production reaches a plateau already at micromolar concentrations (i.e., far below the concentrations at which KIC induces insulin release), it is proposed here that the catabolism of KIC may induce additional signals related to insulin release.
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PMID:Alpha-ketoisocaproate is not a true substrate for ATP production by pancreatic beta-cell mitochondria. 951 37


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