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)

Thermococcus litoralis is a strictly anaerobic archaeon that grows at temperatures up to 98 degrees C by fermenting peptides. Little is known about the primary metabolic pathways of this organism and, in particular, the role of enzymes that are dependent on thermolabile nicotinamide nucleotides. In this paper we show that the cytoplasmic fraction of cell extracts contained NADP-specific glutamate dehydrogenase (GDH) and NADP-specific alcohol dehydrogenase (ADH) activities, neither of which utilized NAD as a cofactor. The GDH is composed of identical subunits having an M(r) of 45,000 and had an optimal pH and optimal temperature for glutamate oxidation of 8.0 and > 95 degrees C, respectively. Potassium phosphate (60 mM), KCl (300 mM), and NaCl (300 mM) each stimulated the rate of glutamate oxidation activity between two- and threefold. For glutamate oxidation the apparent Km values at 80 degrees C for glutamate and NADP were 0.22 and 0.029 mM, respectively, and for 2-ketoglutarate reduction the apparent Km values for 2-ketoglutarate, NADPH, and NH4+ were 0.16, 0.14, and 0.63 mM, respectively. This enzyme is the first NADP-specific GDH purified form a hyperthermophilic organism. T. litoralis ADH is a tetrameric protein composed of identical subunits having an M(r) of 48,000; the optimal pH and optimal temperature for ethanol oxidation were 8.8 and 80 degrees C, respectively. In contrast to GDH activity, potassium phosphate (60 mM), KCl (0.1 M), and NaCl (0.3 M) inhibited ADH activity, whereas (NH4)2SO4 (0.1 M) had a slight stimulating effect. This enzyme exhibited broad substrate specificity for primary alcohols, but secondary alcohols were not oxidized.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Purification and characterization of NADP-specific alcohol dehydrogenase and glutamate dehydrogenase from the hyperthermophilic archaeon Thermococcus litoralis. 813 16

Pig kidney dopa decarboxylase (DDC) expressed in Escherichia coli is a homodimeric enzyme containing one catalytically active pyridoxal 5'-phosphate active site per subunit. In addition to catalyzing the decarboxylation of -aromatic amino acids, DDC also reacts with 5-hydroxytryptamine (5-HT), converting it to 5-hydroxyindolacetaldehyde and ammonia. These products have been identified by means of the enzymes alcohol dehydrogenase and glutamate dehydrogenase, together with high performance liquid chromatographic and mass spectroscopic analysis. The Kcat and Km values of this reaction were determined to be 0.48 min-1 and 0.47 mM, respectively. The NaBH4-reduced enzyme does not catalyze this reaction. Concurrent with this reaction, 5-HT inactivates DDC in both a time- and concentration-dependent manner and exhibits saturation of the rate of inactivation at high concentrations, with Ki and Kinact values of 0.40 mM and 0.023 min-1, respectively. Protection from inactivation by 5-HT was observed in the presence of the active site-directed inhibitor 3,4-dihydroxy-D-phenylalanine. Inactivation with [2-14C]5-HT results in the incorporation of 1 mol of label/enzyme subunit. Taken together, these findings indicate that 5-HT is both a substrate and a mechanism-based inactivator with a partition ratio for product formation versus inactivation of 21. The absorbance, CD, and fluorometric features of 5-HT-inactivated DDC have also been characterized. A speculative mechanism for the reaction and inactivation consistent with the experimental findings is presented.
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PMID:Mechanism-based inactivation of dopa decarboxylase by serotonin. 879 28

The main pathway for the hepatic oxidation of ethanol to acetaldehyde proceeds via ADH and is associated with the reduction of NAD to NADH; the latter produces a striking redox change with various associated metabolic disorders. NADH also inhibits xanthine dehydrogenase activity, resulting in a shift of purine oxidation to xanthine oxidase, thereby promoting the generation of oxygen-free radical species. NADH also supports microsomal oxidations, including that of ethanol, in part via transhydrogenation to NADPH. In addition to the classic alcohol dehydrogenase pathway, ethanol can also be reduced by an accessory but inducible microsomal ethanoloxidizing system. This induction is associated with proliferation of the endoplasmic reticulum, both in experimental animals and in humans, and is accompanied by increased oxidation of NADPH with resulting H2O2 generation. There is also a concomitant 4- to 10-fold induction of cytochrome P4502E1 (2E1) both in rats and in humans, with hepatic perivenular preponderance. This 2E1 induction contributes to the well-known lipid peroxidation associated with alcoholic liver injury, as demonstrated by increased rates of superoxide radical production and lipid peroxidation correlating with the amount of 2E1 in liver microsomal preparations and the inhibition of lipid peroxidation in liver microsomes by antibodies against 2E1 in control and ethanol-fed rats. Indeed, 2E1 is rather "leaky" and its operation results in a significant release of free radicals. In addition, induction of this microsomal system results in enhanced acetaldehyde production, which in turn impairs defense systems against oxidative stress. For instance, it decreases GSH by various mechanisms, including binding to cysteine or by provoking its leakage out of the mitochondria and of the cell. Hepatic GSH depletion after chronic alcohol consumption was shown both in experimental animals and in humans. Alcohol-induced increased GSH turnover was demonstrated indirectly by a rise in alpha-amino-n-butyric acid in rats and baboons and in volunteers given alcohol. The ultimate precursor of cysteine (one of the three amino acids of GSH) is methionine. Methionine, however, must be first activated to S-adenosylmethionine by an enzyme which is depressed by alcoholic liver disease. This block can be bypassed by SAMe administration which restores hepatic SAMe levels and attenuates parameters of ethanol-induced liver injury significantly such as the increase in circulating transaminases, mitochondrial lesions, and leakage of mitochondrial enzymes (e.g., glutamic dehydrogenase) into the bloodstream. SAMe also contributes to the methylation of phosphatidylethanolamine to phosphatidylcholine. The methyltransferase involved is strikingly depressed by alcohol consumption, but this can be corrected, and hepatic phosphatidylcholine levels restored, by the administration of a mixture of polyunsaturated phospholipids (polyenylphosphatidylcholine). In addition, PPC provided total protection against alcohol-induced septal fibrosis and cirrhosis in the baboon and it abolished an associated twofold rise in hepatic F2-isoprostanes, a product of lipid peroxidation. A similar effect was observed in rats given CCl4. Thus, PPC prevented CCl4- and alcohol-induced lipid peroxidation in rats and baboons, respectively, while it attenuated the associated liver injury. Similar studies are ongoing in humans.
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PMID:Role of oxidative stress and antioxidant therapy in alcoholic and nonalcoholic liver diseases. 889 26

Previous studies have capitalized on ordered kinetic mechanisms in the design of biospecific affinity chromatographic methods for highly efficient purifications and mechanistic studies of enzymes. The most direct tactic has been the use of immobilised analogues of the following, usually enzyme-specific substrates, e.g., lactate/pyruvate in the case of lactate dehydrogenase for which NAD+ is the leading substrate. Such immobilised specific substrates are, however, often difficult or impossible to synthesise. The locking-on strategy reverses the tactic by using the more accessible immobilised leading substrate, immobilised NAD+, as adsorbent with soluble analogues of the enzyme-specific ligands (e.g., lactate in the case of lactate dehydrogenase) providing a substantial reinforcement of biospecific adsorption sufficient to effect adsorptive selection of an enzyme from a group of enzymes such as the NAD(+)-specific enzymes. The value of this approach is demonstrated using model studies with lactate dehydrogenase (LDH, EC 1.1.1.27), alcohol dehydrogenase (ADH, EC 1.1.1.1), glutamate dehydrogenase (GDH, EC 1.4.1.3) and malate dehydrogenase (MDH, EC 1.1.1.37). Purification of bovine liver GDH in high yield from crude extracts is described using the tactic.
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PMID:Further studies on the bioaffinity chromatography of NAD(+)-dependent dehydrogenases using the locking-on effect. 891 27

This report enquires on the potentiality of Trp phosphorescence for probing the conformational state of proteins deposited on solid dry films. Thin, amorphous protein films were fabricated with Apoazurin, alcohol dehydrogenase, glyceraldehyde-3-phosphate dehydrogenase and glutamate dehydrogenase the protein being incorporated into a DEAE-dextran matrix and deposited on quartz slides. The results, obtained with appositely constructed instrumentation, demonstrate that thanks to the low background radiation associated with long-lived, delayed emission phosphorescence can be readily detected down to single protein layer matrices and that both spectrum and lifetime are important indicators of the integrity of the protein globular fold. In fact, denaturation of the proteins by guanidinium hydrochloride or heat treatment points out that disruption of the native fold leads to a red shift and broadening of the spectrum with loss of vibronic structure, accompanied to considerably shorter-lived and more heterogeneous decay kinetics. It is also shown that the sensitivity of the phosphorescence lifetime towards the detection of altered, looser conformations of the polypeptide are remarkably enhanced on partial hydration of the sample.
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PMID:Tryptophan phosphorescence as a monitor of protein conformation in molecular films. 1141 43

Twelve enzymes from mature pollen grains of maize were separated by two-dimensional polyacrylamide gel electrophoresis (2-D PAGE). The separation in the second dimension was both in the presence and absence of sodium dodecyl sulfate (SDS). Ten of the investigated enzymes lost activity after separation in the presence of SDS, but those of esterases and acid phosphatase could be recovered. On the other hand, 2-D electrophoresis without SDS is suitable for the analysis of maize pollen pectinesterase, malate dehydrogenase, glutamic-oxalacetic transaminase, diaphorase, superoxide dismutase, and phosphoglucose isomerase. 1-D PAGE and isoelectric focusing (IEF) are sufficient to analyze glucose-6-phosphate dehydrogenase, alcohol dehydrogenase, shikimic dehydrogenase, and glutamate dehydrogenase. The possibility of applying 2-D electrophoresis for the analysis of enzymes from single stigma and stigma exudate is dicussed.
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PMID:Maize pollen enzymes after two-dimensional polyacrylamide gel electrophoresis in the presence or absence of sodium dodecyl sulfate. 1182 13

A series of molecular species with approximately spherical shape and with molecular weights between 35,000 and 250,000 were shadowed with platinum while resting on a cleaved mica surface. They were backed, stripped from the surface, and examined by electron microscopy. Materials examined were: pepsin, liver alcohol dehydrogenase, yeast alcohol dehydrogenase, glutamic dehydrogenase, polyhedral virus protein (insect), fibrinogen substructure, alkaline phosphatase, and microsomal particles from Escherichia coli. Measurements were made of widths perpendicular to the shadowing direction and heights were deduced from shadow lengths. For those molecular species with well established molecular weights the average heights correlate very well with the diameter of the theoretical sphere but the average widths are too great by 50 to 80 A due to the lateral growth of the deposited metal. Although the distortion in shape of shadowed particles is relatively large, with standardized conditions for shadowing, it is possible to make allowance for the distortion and to obtain reasonably reliable estimates of the dimensions of spherical organic particles down to a molecular weight of about 35,000.
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PMID:Measurement of globular protein molecules by electron microscopy. 1439 16

The present study evaluates the expression of genes of Giardia lamblia, one of the most simple and most early diverging eukaryotes, that encode the metabolic enzymes pyruvate: ferredoxin oxidoreductase (PFOR), acetyl-CoA synthetase (ACS), alcohol dehydrogenase E (ADHE) and glutamate dehydrogenase (GDH) and the cyst wall protein (CWP1) gene in trophozoites, cysts and during the excystation process. Primers were designed to amplify mRNA fragments through quantitative reverse-transcriptase-polymerase-chain-reaction. In trophozoites, all transcripts of the enzymes studied were present. In cysts, three of the transcripts were detected: CWP1, GDH and ACS; but the relative levels of the mRNA of GDH and ACS were very different between trophozoites and cysts. During excystation, PFOR and ADHE transcripts appeared after the first induction phase, and the mRNAs of ACS and GDH increased throughout the process.
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PMID:Transcription of metabolic enzyme genes during the excystation of Giardia lamblia. 1466 85

In various populations of the cultivated and weedy amaranth species, the electrophoretic patterns of alcohol dehydrogenase (ADH), glutamate dehydrogenase (GDH), malate dehydrogenase (MDH), isocitrate dehydrogenase (IDH) and malic enzyme (Me) were studied. In total, 52 populations and two varieties (Cherginskii and Valentina) have been examined. Allozyme variation of this material was low. Irrespective of species affiliation, 26 populations and two varieties were monomorphic for five enzymes; a slight polymorphism of three, two, and one enzymes was revealed in three, nine, and fourteen populations, respectively. A single amaranth locus, Adh, with two alleles, Adh F and Adh S, controls amaranth ADH. Two alleles, common Gdh S and rare Gdh F, control GDH; no heterozygotes at this locus were found. The MDH pattern has two, the fast- and slow-migrating, zones of activity (I and II, respectively). Under the given electrophoresis conditions, the fast zone is diffuse, whereas slow zone is controlled by two nonallelic genes, monomorphic Mdh 1 and polymorphic Mdh 2 that includes three alleles: Mdh 2-F, Mdh 2-N, and Mdh 2-S. Low polymorphism of IDH and Me was also found, though their genetic control remains unknown.
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PMID:[Isozyme analysis in a genetic collection of amaranths (Amaranthus L)]. 1639 55

Isozyme phenotypes were determined for 101 strains of Gibberella fujikuroi and 2 strains of Gibberella nygamai that represent seven biological species (mating populations) isolated from a variety of plant hosts in dispersed geographic locations. Fourteen enzymes were resolved in one or more of three buffer systems. Two of the enzymes, arylesterase and acid phosphatase, were polymorphic within two or more biological species and are suitable for intraspecific studies of population variation. Six enzymes, alcohol dehydrogenase, aspartate aminotransferase, glucose-6-phosphate dehydrogenase, mannitol dehydrogenase, phosphoglucomutase, and phosphogluconate dehydrogenase, were monomorphic in all of the isolates examined. The remaining six enzymes, fumarase, glucose phosphate isomerase, glutamate dehydrogenase (NADP), isocitrate dehydrogenase (NADP), malate dehydrogenase, and triose-phosphate isomerase, could potentially be used to distinguish the different biological species. Mating populations C and D are the most similar, since the mating population C isolates examined had the same isozyme phenotype as did a subset of the isolates in mating population D. Mating population E is the least similar to the other taxa examined. Unique isozyme phenotypes are present but are composed of banding patterns shared among the biological species. This finding supports the hypothesis that these biological species, with the possible exception of mating populations C and D, are reproductively isolated from one another and that no significant gene flow is occurring between them. Isozyme analysis is a useful method to distinguish these closely related biological species. Examination of isozyme phenotypes is more rapid than the present technique, which is based on sexual crosses; can be applied to strains that are not sexually fertile; and is more sensitive than traditional morphological characters, which cannot distinguish more than three or four morphological groups among the seven biological species. While emphasizing the discreteness of the mating populations as biological entities, our isozyme data also reaffirm the close genetic relationship among these groups.
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PMID:Isozyme Variation among Biological Species in the Gibberella fujikuroi Species Complex (Fusarium Section Liseola). 1653 23


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