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)

The maximal rates (Vmax) of some enzyme activities related to synaptosomal energy metabolism were studied in different types of synaptosomes from cerebellar cortex of Macaca Fascicularis (Cynomolgus monkey). Different synaptosomal populations, namely "large" and "small" synaptosomes, were isolated from the anterior lobule of the cerebellar cortex of monkeys treated p.o. with dihydroergocriptine at the dose of 12 mg/kg/day before and during the induction of a Parkinson's-like syndrome by MPTP administration (i.v., 0.3 mg/kg/day for 5 days). The enzymes were chosen according to their regulatory role and as markers of the following metabolic pathways: (a) glycolysis ((hexokinase, phosphofructokinase, lactate dehydrogenase), (b) Krebs' (TCA) cycle (citrate synthase, malate dehydrogenase), (c) amino acid, glutamate metabolism (glutamate dehydrogenase, glutamate-pyruvate- and glutamate-oxaloacetate-transaminases), (d) acetylcholine catabolism (acetylcholinesterase) and (e) ATPases, i.e. Na(+)-K(+)-ATPase, Mg(2+)-ATP synthetase, Mg(2+)-ATPase, Ca(2+)-Mg(2+)-ATPase and Ca(2+)-ATPase Low and High affinity for Ca2+. The MPTP administration modified the activities of citrate synthase, malate dehydrogenase, Na(+)-K(+)-ATPase, acetylcholinesterase and glutamate-oxaloacetate transaminase only on selected types of synaptosomes. Pharmacological treatment by dihydroergocriptine was able to recovery at the steady-state levels the activities of these enzymes, thus demonstrating a partial protective effect on these biochemical parameters.
...
PMID:Parkinson-like disease by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) toxicity in Macaca fascicularis: synaptosomal metabolism and action of dihydroergocriptine. 817 63

We previously identified a 50 kDa membrane protein which bound to in vitro assembled microtubules [Mithieux and Rousset (1989) J. Biol. Chem. 264, 4664-4668]. This protein exhibited the expected properties for mediating the ATP-dependent association of vesicles with microtubules [Mithieux, Audebet and Rousset (1988) Biochim. Biophys. Acta 969, 121-130]. The 50 kDa membrane protein (MP50), initially extracted in very low amount from isolated pig thyroid lysosomes/endosomes, has now been purified from membrane preparations of crude vesicle fractions from pig liver and brain. MP50 was isolated from detergent-solubilized membrane protein by affinity chromatography on immobilized ATP; 3-5 mg of MP50 was obtained from 100 g of liver tissue. Phase partitioning in Triton X-114 indicated that MP50 is a peripheral membrane protein. Radioiodinated liver MP50 bound to microtubules assembled in vitro. The binding was inhibited by ATP (Ki = 0.76 mM) and displaced by unlabelled liver or brain MP50. Equilibrium binding studies yielded KD values of 1.8 x 10(-7) M. By N-terminal amino acid sequence analysis, MP50 was identified as glutamate dehydrogenase (GDH), by comparison of V8 protease peptide maps of MP50 with purified liver GDH. Liver MP50 exhibited a low GDH activity; 4-5 units/mg compared with 18 and 34 units/mg for purified bovine and rat liver GDH respectively. Bovine and rat liver GDH yielded six spots from pI 5.7 to 7.2 when analysed by two-dimensional electrophoresis; in contrast, MP50 gave one main spot (corresponding to spot 2 of liver GDH) with a pI of approx. 6.5. Soluble liver GDH from commercial sources exhibited a very low or no microtubule-binding activity. In conclusion, we have found a membrane-bound form of GDH capable of specific and nucleotide-sensitive interaction with microtubules. Our data suggest that GDH isoproteins, the number of which has been undervalued up to now, could have cellular functions other than that of an enzyme.
...
PMID:A membrane-bound form of glutamate dehydrogenase possesses an ATP-dependent high-affinity microtubule-binding activity. 824 Feb 42

The energy metabolism of a mammalian cell line grown in vitro was analyzed by substrate consumption rates and metabolic flux measurements. The data allowed the determination of the relative importance of the pathways of glucose and glutamine metabolism to the energy requirements of the cell. Changes in the substrate concentrations during culture contributed to the changing catalytic activities of key enzymes, which were determined. 1. A murine B-lymphocyte hybridoma (PQXB1/2) was grown in batch culture to a maximum cell density of 1-2 x 10(6) cells/mL in 3-4 d. The intracellular protein content showed a maximum value during the exponential growth phase of 0.55 mg/10(6) cells. Glutamine was completely depleted, but glucose only partially depleted to 50% of its original concentration when the cells reached a stationary phase following exponential growth. 2. The specific rates of glutamine and glucose utilization varied during culture and showed maximal values at the midexponential phase of 2.4 nmol/min/10(6) cells and 4.3 nmol/min/10(6) cells, respectively. 3. A high proportion of glucose (96%) was metabolized by glycolysis, but only limited amounts by the pentose phosphate pathway (3.3%) and TCA cycle (0.21%). 4. The maximum catalytic activity of hexokinase approximates to the measured flux of glycolysis and is suggested as a rate-limiting step. In the stationary phase, the hexokinase activity reduced to 11% of its original value and may explain the reduced glucose utilization at this stage. 5. The maximal activities of two TCA cycle enzymes were well above the measured metabolic flux and are unlikely to pose regulatory barriers. However, the activity of pyruvate dehydrogenase was undetectable by spectrophotometric assay and explains the low level of flux of glycolytic metabolites into the TCA cycle. 6. A significant proportion of the glutamine (36%) utilized by the cells was completely oxidized to CO2. 7. The measured rate of glutamine transport into the cells approximated to the metabolic flux and is suggested as a rate-limiting step. 8. Glutamine metabolism is likely to occur via glutaminase and amino transaminase, which have significantly higher activities than glutamate dehydrogenase. 9. The calculated potential ATP production suggests that, overall, glutamine is the major contributor of cellular energy. However, at the midexponential phase, the energy contribution from the catabolism of the two substrates was finely balanced--glutamine (55%) and glucose (45%).
...
PMID:Glucose and glutamine metabolism of a murine B-lymphocyte hybridoma grown in batch culture. 826 5

Mature mitochondrial proteins (aspartate aminotransferase, malate dehydrogenase, hydroxyacyl coenzyme A dehydrogenase, creatine kinase) and cytosolic proteins (aldolase, glyceraldehyde-3-phosphate dehydrogenase) with a basic pI were found to bind to isolated mitochondria, electrostatic interactions being mainly responsible for their binding. Mitochondrial aspartate aminotransferase bound with a Kd' of 30 nM in 0.6 M sorbitol, 20 mM Hepes/KOH, pH 7.4, at 25 degrees C. Cytosolic aspartate aminotransferase and glutamate dehydrogenase (a protein located in the mitochondrial matrix) both with an acidic pI, did not bind to mitochondria. Treatment of mitochondria with proteinases did not affect the subsequent binding of imported mitochondrial proteins. Their association with both intact and proteinase-treated mitochondria resulted in a marked increase in their susceptibility toward proteinase K. In contrast, the basic cytosolic proteins tested bound only to intact mitochondria and thereby did not become more susceptible toward proteolytic attack. Treatment of mitochondria with adriamycin, a drug binding to acidic phospholipids, prevented the subsequent association of mitochondrial aspartate aminotransferase with mitochondria and the ensuing conformational labilization. Apparently, the mature moiety of imported mitochondrial proteins is partially unfolded upon interaction with the lipid component of the mitochondrial envelope. Both the binding of the mitochondrial proteins and their conformational labilization is independent of ATP and the electrochemical potential across the inner membrane.
...
PMID:The mature form of imported mitochondrial proteins undergoes conformational changes upon binding to isolated mitochondria. 828 42

Two "targeted bidentate" photoaffinity cross-linking reagents, the monoanhydride of 8-N3ADP with N-(4-(benzoyl)phenylmethyl)phosphoramide ([gamma-32P]8-N3ATP gamma BP) and the monoanhydride of 8-N3GDP with N-(4-(benzoyl)phenylmethyl)-phosphoramide ([gamma-32P]8-N3GTP gamma BP), were developed for studying the inter- and intramolecular interactions of nucleotide-binding proteins. Experiments using these bidentate reagents with two photoactive groups led to specific cross-linking: [gamma-32P]8-N3GTP gamma BP and [gamma-32P]8-N3ATP gamma BP showed intersubunit cross-linking of glutamate dehydrogenase and [gamma-32P]8-N3GTP gamma BP appeared to cross-link the alpha- and beta-subunits of tubulin. The non-azido "monodentate" versions of these reagents, the monoanhydride of ADP with N-(4-(benzoyl)phenylmethyl)-phosphoramide ([gamma-32P]ATP gamma BP) and the monoanhydride of GDP with N-(4-(benzoyl)phenylmethyl)-phosphoramide ([gamma-32P]GTP gamma BP), were also synthesized and characterized. The ability of these monodentate reagents with one photoactive group to serve as photoaffinity probes was established by photolabeling specifically the exchangeable GTP-binding domain of tubulin with [gamma-32P]GTP gamma BP and the ATP-binding domain of purified adenylate kinase and several nucleotide-binding proteins in human brain homogenate with [gamma-32P]ATP gamma BP.
...
PMID:Synthesis and application of bidentate photoaffinity cross-linking reagents. Nucleotide photoaffinity probes with two photoactive groups. 831 86

This study was undertaken in order to assess the role of purely circulation-related effects upon free-radical-mediated reperfusion injury in the liver by comparing the respective effects of the oxygen free-radical scavenger superoxide dismutase (SOD) and the vasodilative action of papaverine in an ischemia/reperfusion model of the liver. Livers from male Wistar rats were rinsed blood free via the portal vein and stored ischemically (60 min at 37 degrees C in Krebs-Henseleit solution and 60 min at 4 degrees C in Euro-Collins solution). Reperfusion was carried out at a constant flow of 30 ml/min for 45 min at 37 degrees C in a nonrecirculating manner. Warm ischemic damage was evident in untreated livers compared to control livers, submitted solely to cold ischemia for 2 h at 4 degrees C, by increased vascular resistance upon reperfusion, enhanced enzyme leakage from the parenchyma (glutamate pyruvate transaminase, glutamate dehydrogenase) and from the endothelium (purine-nucleoside phosphorylase), reduced tissue content of ATP and enhanced lipid peroxidation. Preischemic treatment with SOD or papaverine (the latter also given during reperfusion) significantly reduced hepatic vascular resistance and parenchymal enzyme loss in a comparable manner. Both drugs resulted in a significant increase of hepatic tissue content of ATP at the end of reperfusion. SOD, but not papaverine, prevented the leakage of purine-nucleoside phosphorylase and significantly reduced the tissue levels of lipid peroxides.(ABSTRACT TRUNCATED AT 250 WORDS)
...
PMID:Role of the hepatovasculature in free radical mediated reperfusion damage of the liver. 840 87

Photoaffinity labeling with [alpha-32P]8N3GTP and [gamma-32P]8N3GTP was used to identify the guanine binding domain of the GTP regulatory site within glutamate dehydrogenase (GDH). Without photolysis, 8N3GTP mimicked the regulatory properties of GTP on GDH activity with 8N3GTP exhibiting a Ki of 5 microM while the Ki for GTP was about 0.6 microM. Under optimal photolabeling conditions saturation of photoinsertion with 1 microgram of GDH revealed an apparent Kd of 9 +/- 4 microM for [gamma-32P]8N3GTP. Photolabeling with this analog could be competitively inhibited with GTP with an apparent Kd of 12 +/- 2 microM. Other nucleotides such as ATP and NAD(P)H could not reduce the amount of photoinsertion as effectively as GTP. ADP could decrease photoinsertion, but only at much higher concentrations. NAD(P)+, GDP, AMP, and GMP had little effect on photoinsertion. Divalent cations Mg2+ and Ca2+ also reduced photoinsertion significantly while the monovalent K+ and Na+ ions had no effect. Aluminum(III)-chelate or iron(III)-chelate affinity chromatography and reversed-phase HPLC were used to purify photolabel-containing peptides generated with either trypsin or chymotrypsin. This identified a portion of the guanine binding domain within the GTP regulatory site as the region containing the sequence Ile439 to Tyr454. Photolabeling of this peptide was prevented 91% by the presence of 300 microM GTP during photolysis. Lys445 was not identified in sequence analyses of the photolabeled peptides. Also, trypsin was unable to cleave the photolabeled peptide at this site. These results suggest that Lys445 may be the residue modified by [alpha-32P]8N3GTP.
...
PMID:Identification of a guanine binding domain peptide of the GTP binding site of glutamate dehydrogenase: isolation with metal-chelate affinity chromatography. 843 45

A variety of metabolites have been found to elicit a form of inhibition or activation on an NAD-specific glutamate dehydrogenase (NAD-GDH, EC 1.4.1.2) from Halobacterium halobium. The purified halophilic enzyme was tested with several compounds known to be allosteric modifiers of mammalian glutamate dehydrogenases to determine their effects on enzyme activity. GTP, ATP, ADP and AMP did not affect the enzyme, so these effectors of bovine glutamate dehydrogenase do not play a role in the regulation of the halophilic enzyme. However, the halophilic enzyme was subject to strong inhibition by TCA intermediates. When measuring the initial rate of the reaction, the oxidative deamination of L-glutamate was inhibited by TCA metabolites such as: fumarate, oxalacetate, succinate and malate; by substrate analogues such as: NADP+, D-glutamate and glutarate; and by dicarboxylic compounds such as adipate. On the other hand, all the amino acids tested were activators of this enzyme, except the D-isomer of the substrate L-glutamate that acted as an inhibitor. The relative effectiveness of each inhibitor or activator (Ki or Ka values) was correlated with the dipole moment (mu), HOMO and LUMO molecular orbital energies, optimal distance between two carboxyl groups, and hydrophobicity. Compounds with high dipole moment acted as good activators while compounds with low dipole moment were inhibitors. We have also found that the best activators were amino acids with no polar lateral chain.
...
PMID:NAD-glutamate dehydrogenase from Halobacterium halobium: inhibition and activation by TCA intermediates and amino acids. 860 24

A one-step perifusion technique is described for studying the regulation of energy metabolism in intact hepatocytes and in mitochondria of the same cells after their permeabilization by digitonin. Cell count and activities of glutamate dehydrogenase, the latter being used as an indicator of mitochondrial integrity, were found to be nearly unchanged after permeabilization and perifusion for at least 40 min at 37 degrees C. The residual activity of lactate dehydrogenase after permeabilized indicated that permeabilized cells were almost depleted of soluble cytosolic components. The composition of the perifusion medium was chosen so that various metabolic states could be adjusted of both intact and permeabilized hepatocytes without the need to change the perfusion medium. Oxidative phosphorylation of mitochondria within permeabilized hepatocytes remained intact throughout the perifusion as indicated by the response of respiration to the addition of ADP, carboxyatractyloside and uncoupler. The application of the perifusion technique allows us to sample indicator metabolites in the effluent medium like acetoacetate (AcAc) and 3-hydroxybutyrate (HB) for calculating the mitochondrial redox ratios and rates of ketogenesis. In the presence of octanoate and ADP, an improvement of substrate supply by glutamate and malate led to increases in the intramitochondrial HB/AcAc ratio and the respiration rate. Glutamate/malate concentrations of 1 mM resulted in maximal respiration rates, whereas concentrations of 5 mM further enhanced the HB/AcAc ratio. Mitochondria responded to increasing ATP/ADP ratios in the perifusion medium by decreased respiration rates at higher HB/AcAc ratios. By comparing respiration rates and redox ratios of mitochondria in permeabilized cells with those before permeabilization (gluconeogenic conditions of hepatocytes), it is concluded that in the intact cell oxidative phosphorylation is limited with respect to substrate supply as well as by the ATP demand.
...
PMID:Interrelation between mitochondrial respiration, substrate supply and redox ratio in perifused permeabilized rat hepatocytes. 861 60

We established a simple and rapid kinetic assay for measurement of calcium in serum by using urea amidolyase (EC 3.5.1.45) from yeast species. The method is based on inhibition of the enzyme by calcium. In the assay, we eliminated endogenous ammonium ion by use of glutamate dehydrogenase (GLDH; EC 1.4.1.4); then in the presence of urea amidolyase, urea, ATP, bicarbonate, magnesium, and potassium ions, ammonium ion production was inversely proportional to calcium ion concentration in serum. The concentration of ammonium ion formed was determined by adding GLDH to produce NADP+ in the presence of 2-oxoglutarate and NADPH; we then monitored the change of absorbance at 340 nm. The within-run CVs of this method were 1.7-3.2% (n = 10) at 1.53-3.08 mmol/L, respectively. Day-to-day (total) CVs were 2.8-4.1%. Analytical recovery was 92-112%. The presence of other ions, ascorbic acid, reduced glutathione, bilirubin, hemoglobin, citrate, lipemic material, or human serum albumin did not affect this assay system. The correlation between values obtained with our method (y) and o-cresolphthalein complexone method (CPC) (x) was: y = 1.001x + 0.077 mmol/L (r = 0.949, Sy[symbol: see text]x = 0.079, n = 100); with the other enzymatic method (x) it was: y = 0.952x + 0.021 mmol/L (r = 0.955, Sy[symbol: see text]x = 0.074, n = 100). The SEs for each method were: 0.025 mmol/L, our method; 0.023 mmol/L, CPC method; and 0.025 mmol/L, the other enzymatic method.
...
PMID:New enzymatic assay for calcium in serum. 869 77


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

---