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:1.4.3.11 (glutamate dehydrogenase)
4,437 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. A polarographic assay of superoxide (O2--) dismutase (EC 1.15.1.1) activity is described, in which the ability of the enzyme to inhibit O2---dependent sulphite oxidation, initiated by xanthine oxidase activity, is measured. The assay was used in a study of the intracellular distribution of superoxide dismutase in rat liver. Both cyanide-sensitive cupro-zinc dismutase (92% of the total activity) and cyanide-insensitive mangano-dismutase (8%) were measured. 2. Rat liver homogenates contained both particulate (16%y and soluble (84%) dismutase activity. The particulate activity contained both types of dismutase, whereas nearly all the soluble dismutase was a cupro-zinc enzymes. The distribution pattern of mangano-dismutase was similar to that of cytochrome oxidase and glutamate dehydrogenase, indicating that the enzyme was probably present exclusively in the mitochondria. 3. Superoxide dismutase activity in the heavy-mitochondrial (M) fraction was latent and was activated severalfold and largely solubilized by sonication. Treatment of the M fraction with digitonin or a hypo-osmotic suspending medium indicated that most of the cupro-zinc dismutase was located in the mitochondrial intermembrane space, whereas the mangano-enzyme was located in the inner-membrane and matrix space. 4. A small amount of dismutase activity appeared to be present in the nuclei and microsomal fraction, but little or no activity in the lysosomes or peroxisomes. 5. The results are discussed in relation to the intracellular location of known O2---generating enzymes, the possible role of superoxide dismutase activity in intracellular H2O2 formation, and to current views on the physiological function of the enzyme.
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PMID:Polarographic assay and intracellular distribution of superoxide dismutase in rat liver. 81 Jan 38

The catalytic activity of the enzyme L-glutamic acid decarboxylase (GAD) is determined by an amperometric method based on a recently developed glutamate-selective biosensor. The biosensor is composed of an amperometric H2O2 electrode and a biocatalytic membrane containing the enzyme glutamic acid oxidase (GAO). The biosensor allows the direct and continuous measurement of GA levels by monitoring the H2O2 produced at the electrode interface as a coproduct of the GAO-catalyzed GA oxidation to alpha-ketoglutaric acid. Since GA is transformed to gamma-aminobutyric acid and CO2 under the catalytic activity of GAD, the rate of GA consumption in solution, monitored by the GAO biosensor, represents a reliable measure of GAD catalytic activity. Additional experiments performed in the presence of different concentrations of the GAD inhibitor valproic acid have shown the suitability of the proposed approach for the study of GAD inhibitors also. Discussion of the main experimental characteristics of this new analytical method is given in terms of sensitivity, reproducibility, and reliability of the experimental results and ease, time, and cost of operation.
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PMID:Determination of glutamic acid decarboxylase activity and inhibition by an H2O2-sensing glutamic acid oxidase biosensor. 135 47

The glutamine synthetase and the NADP-specific glutamate dehydrogenase activities of Neurospora crassa were lost in a culture without carbon source only when in the presence of air. Glutamine synthetase was previously reported to be liable to in vitro and in vivo inactivation by activated oxygen species. Here we report that NADP-specific glutamate dehydrogenase was remarkably stable in the presence of activated oxygen species but was rendered susceptible to oxidative inactivation when chelated iron was bound to the enzyme and either ascorbate or H2O2 reacted on the bound iron. This reaction gave rise to further modifications of the enzyme monomers by activated oxygen species, to partial dissociation of the oligomeric structure, and to precipitation and fragmentation of the enzyme. The in vitro oxidation reaction was affected by pH, temperature, and binding to the enzyme of NADPH. Heterogeneity in total charge was observed in the purified and immunoprecipitated enzymes, and the relative amounts of enzyme monomers with different isoelectric points changes with time of the oxidizing reaction.
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PMID:Oxidation of Neurospora crassa NADP-specific glutamate dehydrogenase by activated oxygen species. 253 Feb 8

A chemiluminometric method for the automated flow injection analysis of ammonia is described. The essence of the invention is the use of a bioreactor consisting of both immobilized L-glutamate dehydrogenase (GLDH) and L-glutamate oxidase (GLXD), which are sequentially aligned in this order in a minicolumn measuring 2.0 X 20 mm. The unidirectional constant flow of liquid through the column reactor minimizes the reversed diffusion of the solutes so that the following sequence of reactions is ensured. Thus, ammonia to be determined is first transformed by GLDH into L-glutamate, which then produces hydrogen peroxide by GLXD. Hydrogen peroxide in the effluent from the column is then determined by its chemiluminescence upon admixing with luminol and potassium ferricyanide. The present method gives linearity of the standard curve for ammonia up to 1.0 mM. It is at least 100 times more sensitive than the conventional method for ammonia assay using ultraviolet absorption measurement.
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PMID:Use of a bioreactor consisting of sequentially aligned L-glutamate dehydrogenase and L-glutamate oxidase for the determination of ammonia by chemiluminescence. 366 33

An on-line tubular electrode system is described which provides real time measurements of glutamate in brain microdialysate. It is based on the enzyme glutamate oxidase (EC 1.4.3.11) and the detection of H2O2 on a platinum electrode at 600 mV vs Ag/AgCl. The enzyme is immobilized in a layer of o-phenylenediamine electropolymerized at 750 mV in a phosphate buffer, pH 7.0. The layer is 10-15 nm thick and enables good enzyme loading and fast response time. The ability of the polymer to block out electroactive interferents like ascorbate and uric acid combined with a preoxidation system run at 600 mV resulted in a virtually interference free glutamate assay with a lower detection limit of 0.3 microM in the presence of physiological levels of interferents, a sensitivity of 4 nA/microM, and a linear region up to 30 microM. The system is optimized for use with a microdialysis probe implanted in the brain and perfused at 2 microL/min. It provides a time resolution of < 1 min and has been tested in vivo.
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PMID:On-line measurement of brain glutamate with an enzyme/polymer-coated tubular electrode. 791 89

Inactivation of D-amino acid oxidase occurred by different mechanisms. The enzyme showed a rapid loss of activity in the presence of micromolar amounts of Cu2+ and Hg2+. It was also sensitive to oxidative inactivation by Fe2+ and H2O2 when both reagents were added in millimolar amounts. When oxidatively inactivated D-amino acid oxidase and a corresponding non-treated control were modified with the sulfhydryl-modifying, fluorescent reagent monobromobimane and subsequently digested with endoproteinase Glu-C, Cys-298 was identified to be a target for oxidative modification according to differences in the known peptide profile of fluorescence intensity. Another reason for the observed loss of enzyme activity in crude extracts was the specific proteolytic digestion of D-amino acid oxidase, which was dependent on the growth phase of the cells used. This cleavage was catalyzed by a serine-type proteinase and was the introductory step for the further complete degradation of the enzyme. In addition, a coenriched 50-kDa protein, identified as NADPH-specific glutamate dehydrogenase, significantly decreased the stability of the D-amino acid oxidase activity. Treatment of apo-D-amino acid oxidase from T. variabilis with monobromobimane resulted in a significantly increased fluorescence of two peptides, neither of which contained any cysteine residue. Thus, an involvement of cysteine residues in binding the FAD coenzyme should be excluded.
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PMID:Studies on the inactivation of the flavoprotein D-amino acid oxidase from Trigonopsis variabilis. 873 70

Herbivorous voles, Microtus arvalis, have characteristics similar to herbivores in that their hepatic glycolytic enzyme activities are relatively low. The effects of a single low dose (100 mg/kg body weight) of streptozotocin (STZ) in voles were studied and the difference in sensitivity to or toxicity of STZ in voles and C57BL/6 mice was compared. In voles which received STZ, the cumulative incidence of glycosuria reached 53% by 4 weeks after administration. The diabetic voles showed marked increases in their blood glucose and plasma free fatty acid concentrations and a significant decrease in plasma immunoreactive insulin concentrations. Their hepatic hexokinase, glucokinase, glutathione peroxidase and glutamate dehydrogenase activities decreased significantly and lesions were widely observed in the liver, kidney and pancreas. The activities of glutathione peroxidase, a scavenger of H2O2, decreased significantly in their liver and pancreas. These changes were not observed in C57BL/6 mice which received STZ. The higher sensitivity to and toxicity of STZ in voles than in mice are considered to be caused by the characteristically low activities of glycolytic enzymes and glutathione peroxidase in the tissues of voles. Voles may be a good model for studying the mechanisms of cytotoxicity by STZ in herbivorous animals.
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PMID:High sensitivity to streptozotocin in herbivorous voles, Microtus arvalis, compared to mice. 873 20

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

A developmental block is induced by phosphate in rat embryos at the late two-cell stage. The present study was designed to examine the energy metabolism of rat two-cell blocked and non-blocked embryos. Enzyme activity was measured in individual embryos by histochemical techniques. The activities of malate dehydrogenase, isocitrate dehydrogenase, lactate dehydrogenase, pyruvate dehydrogenase, glyceraldehyde-3-phosphate dehydrogenase, glutamate dehydrogenase, glucose-6-phosphate dehydrogenase, glucose-6-phosphatase, and phosphorylase did not differ among non-blocked and blocked embryos. However, the activity of succinate dehydrogenase was significantly decreased in blocked embryos compared with non-blocked embryos. In blocked embryos, cytochrome oxidase activity was distributed homogeneously, but was located at the perinuclear region in non-blocked embryos. Active mitochondrial organization was visualized using the fluorescent probe rhodamine 123 and laser scanning confocal microscopy. In both non-blocked and blocked embryos, mitochondria were distributed homogeneously. The concentration of H2O2 measured fluorometrically in embryos cultured without phosphate did not change significantly during the culture period, but decreased in embryos cultured with phosphate. The timing corresponded to the occurrence of the two-cell block. In summary, these results suggest that the developmental block in rat two-cell embryos is induced by disturbance of mitochondrial energy metabolism.
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PMID:Microscopic analysis of enzyme activity, mitochondrial distribution and hydrogen peroxide in two-cell rat embryos. 986 Nov 63

A selective and sensitive chemiluminometric flow sensor for the determination of L-glutamate in serum, based on immobilized oxidases such as glutamate oxidase (GOD), uricase (UC) and peroxidase (POD), is described herein. The principle for the selective chemiluminometric detection for L-glutamate is based on coupled reactions of four sequentially aligned immobilized oxidases, UC/POD/GOD/POD in a flow cell. The immobilized UC was employed to decompose urate, which is one of the major interfering components in serum for a luminol-H2O2 chemiluminescence reaction. The H2O2 produced from the UC reaction readily reacted with reducing components, such as ascorbate and glutathione, and then the excess H2O2 was decomposed by the immobilized POD. L-Glutamate in the sample plug was enzymatically converted to H2O2 with immobilized GOD. Subsequently, the peroxide reacts with luminol on the immobilized POD to produce chemiluminescence, proportional to glutamate concentration. The enzymes were immobilized on tresylated poly(vinyl alcohol beads). The immobilized enzymes were packed into TPFE tube (1.0 mm i.d. x 60 cm), in turn, and used as a flow cell. The sampling rate was 30 h-1. The calibration graph for L-glutamate is linear for 20 nM-5 microM; the detection limit (signal-to-noise = 3) is 10 nM.
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PMID:Flow-through chemiluminescence sensor using immobilized oxidases for the selective determination of L-glutamate in a flow-injection system. 1170 95


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