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)

Streptococcus bovis JB1 cells energized with glucose transported glutamine at a rate of 7 nmol/mg of protein per min at a pH of 5.0 to 7.5; sodium had little effect on the transport rate. Because valinomycin-treated cells loaded with K and diluted into Na (pH 6.5) to create an artificial delta psi took up little glutamine, it appeared that transport was driven by phosphate-bond energy rather than proton motive force. The kinetics of glutamine transport by glucose-energized cells were biphasic, and it appeared that facilitated diffusion was also involved, particularly at high glutamine concentrations. Glucose-depleted cultures took up glutamine and produced ammonia, but the rate of transport per unit of glutamine (V/S) by nonenergized cells was at least 1,000-fold less than the V/S by glucose-energized cells. Glutamine was converted to pyroglutamate and ammonia by a pathway that did not involve a glutaminase reaction or glutamate production. No ammonia production from pyroglutamate was detected. S. bovis was unable to take up glutamate, but intracellular glutamate concentrations were as high as 7 mM. Glutamate was produced from ammonia via a glutamate dehydrogenase reaction. Cells contained high concentrations of 2-oxoglutarate and NADPH that inhibited glutamate deamination and favored glutamate formation. Since the carbon skeleton of glutamine was lost as pyroglutamate, glutamate formation occurred at the expense of glucose. Arginine deamination is often used as a taxonomic tool in classifying streptococci, and it had generally been assumed that other amino acids could not be fermented. To our knowledge, this is the first report of glutamine conversion to pyroglutamate and ammonia in streptococci.
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PMID:Transport of glutamine by Streptococcus bovis and conversion of glutamine to pyroglutamic acid and ammonia. 272 40

The hybridoma, 62H3, which secretes a monoclonal IgG2b with anti-HLA-DR specificity, was expanded in pristane-primed BALB/c mice and the antibody was isolated from the ascitic fluid by affinity chromatography on Protein A-Sepharose. The purified IgG2b antibody was tested by an enzyme immunoassay for antibody activity against a panel of 40 self and non-self antigens. It was found to react strongly with beta-galactosidase, actin, glutamate dehydrogenase, rabbit and human IgG and di- and trinitrophenyl groups; and moderately with tubulin, insulin and phosphorylcholine; but it did not react with various other self and non-self antigens, such as DNA, albumin, keyhole limpet hemocyanin, hen lysozyme and horseradish peroxidase. Fab and Fc fragments were prepared from this IgG2b by papain proteolysis. The Fab fragment possessed the same spectrum of polyreactivities as the native IgG2b, whereas no activity was detected with the Fc fraction. In order to investigate the properties of the antigen binding site, the actin, TNP and rabbit IgG antibody activities were studied in more detail by enzyme immunoassay, Western blot and immunocytochemistry. The monomolecular nature of this multireactivity was confirmed by immunoabsorption analysis. Furthermore, 62H3 monoclonality was also verified by comparative isoelectric focusing and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis with other monospecific antibodies. The dissociation constants (Kd) of antigen-antibody equilibria in solution were measured. The Kd for actin was 1.11 +/- 0.24 x 10(-5) M and the Kd for TNP-BSA was 8.7 +/- 0.51 x 10(-7) M. No interaction with rabbit IgG could be detected in solution. These findings raise the question of the possible implication in autoimmune pathology or in normal physiology of IgG class polyspecific antibodies with solid-phase restricted cross-reactive rheumatoid factor activity.
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PMID:Immunochemical studies of a murine polyreactive IgG2b autoantibody with rheumatoid factor activity. 277 Jul 48

Sheep received a single intragastric dose of 0.5, 1.0, 1.5, or 2.0 mmol F-/kg. Mild signs occurred at 1.5 mmol F-/kg and the animals recovered 2 days later. With the 2.0 mmol F-/kg dose all animals showed dullness, anorexia, and mild diarrhea which decreased from the third day. Dose-related congestion of duodenum, liver, kidney, and lung was observed in all animals. For the two higher doses kidney degeneration and tubular necrosis were associated with glomerular inflammation. Serum fluoride had a dose-related increase and was still significantly elevated on Day 7 for sheep given doses higher than or equal to 1.0 mmol F-/kg. Serum calcium and glucose levels were significantly lowered for all doses on the first day and the decrease was dose-related. In sheep given 2.0 mmol F-/kg total proteins and sodium were significantly lowered, whereas potassium and urea were increased (p less than 0.05); alkaline phosphatase (ALP) and lactic dehydrogenase (LDH) were both lowered (p less than 0.01) on the first day and ALP was still lowered on Day 7. For the highest dose glutamate dehydrogenase (GDH) was increased on Days 1 and 7 and gamma-glutamyl transferase (GGT) was increased on Day 1 and lowered on Day 7. Diuresis was increased for the two higher doses in Day 3 or 4 following dosage. A dose-related increase of daily fluoride excretion occurred for all doses on Day 1 and fluoride excretion was still significantly elevated on Day 7 except for the lowest dose.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Experimental acute sodium fluoride poisoning in sheep: renal, hepatic, and metabolic effects. 286 58

Studies of various parameters of amino acid and catecholamine metabolism in human cerebral cortex have provided a number of biochemical markers that appear to delineate areas of focal epileptic activity. These observations have been consolidated further by investigations of a number of experimental models of epilepsy in animals. In appraising this data, it is important to take into consideration whether the tissue samples were obtained during an actual seizure state or in an interictal period. It is also important when possible to assess the extent of astrogliosis and neuronal loss. Sites of spontaneously active epileptic spiking in the cerebral neocortex have a somewhat different amino acid profile when compared to gray matter obtained from surrounding nonspiking gyri several centimeters away. There is an elevation in glycine content, a relative diminution in taurine, and a trend towards lowered glutamic acid levels. However, the concentrations of the eight amino acids measured appear in both the foci and surround to still be within the general range for normal tissue. Measurements of key enzymes involved in the synthesis and regulation of neurotransmitters provide a complementary method of evaluating functional changes in epileptic brain as they are generally less labile than their substrates. There is a moderate increase in the activity of glutamic acid dehydrogenase, an enzyme that plays an important role in the synthesis of glutamic acid from glucose. In some patients a decrease in glutamic acid decarboxylase has also been reported: this enzyme forms gamma-aminobutyric acid (GABA) from glutamic acid and is thus important for inhibition in the central nervous system. Moreover, there is a striking increase in the activity of tyrosine hydroxylase, the rate-limiting enzyme responsible for catecholamine synthesis. The possibility of a focal abnormality in catecholamine metabolism is reinforced by the simultaneous finding of a relative decrease in the number of alpha-1 postsynaptic receptor sites. An important marker of energy metabolism in neural tissue, Na+,K+-ATPase activity, has also been found to be decreased in actively spiking human cerebral cortex. Data from experimental animal foci produced by topical application of convulsant agents show a consistent drop in glutamic acid tissue content. This can be matched to an efflux of glutamic acid from the cortical surface, which in turn is proportional to the electrographic activity of the spike focus. In addition, there is often also a decrease in taurine and GABA in such foci, as well as an increase in the levels of a number of neutral amino acids.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Amino acid and catecholamine markers of metabolic abnormalities in human focal epilepsy. 287 18

Using a special tetrazolium salt technique, a striking correlation was observed between Na+ concentration of the incubation medium and the activity of glutamate dehydrogenase (GDH) in glutamatergic neuropil areas of the hippocampal formation, cerebellum, and other brain regions. Na+ concentration of 130 to 150 mmol/l caused maximal formazan production. The histochemical enzyme reaction in neuronal perikarya as well as biochemically estimated GDH activity were found to be rather inhibited by such Na+ concentrations. The GDH catalyzed sodium dependent increase in formazan production is discussed to be a consequence of the sodium dependence of glutamate uptake in glutamatergic brain structures supplying the enzyme with substrate.
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PMID:[Enzyme histochemical demonstration of sodium-dependent glutamate uptake in glutamatergic brain structures in the rat]. 287 83

Proteins of known composition and structural characteristics were incubated (1.0 mg/mL) with re-suspended salivary sediment (2.5% v/v) in a lactate-salt medium with an initial pH of 5.2 for two hr at 37 degrees C. Hydrolysis of the proteins was monitored by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Hydrogen ion, amines, and ammonia were measured by use of a combined pH electrode, high-performance liquid chromatography, and glutamate dehydrogenase, respectively. Of the proteins studied, the caseins alpha s1, beta, and kappa and the histones H1 and H3 were extensively hydrolyzed by the salivary-sediment bacteria. The hydrolysis of these proteins was attributed to their relative lack of tertiary (folded) structure. The only amine detected was the polyamine putrescine arising from the catabolism of arginine following the hydrolysis of the arginine-rich histone H3. None of the other proteins extensively hydrolyzed by salivary sediment, although containing arginyl and lysyl residues, served as substrates for putrescine or cadaverine production. Pre-hydrolysis of the arginine-rich histone H3 and poly-L-arginine with trypsin resulted in a marked increase in putrescine produced, suggesting that the salivary-sediment proteolytic activity was not "trypsin-like". Incubation of salivary-sediment bacteria with the caseins and the histone H3 resulted in an increase in ammonium ion concentration and an associated decrease in hydrogen ion concentration. The increase in ammonium ion concentration not attributed to arginine hydrolysis was correlated with the content of glutaminyl plus asparaginyl residues of the proteins.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Protein dissimilation by human salivary-sediment bacteria. 291 33

We describe a procedure using immobilized nicotinamide as an affinity chromatographic ligand for the binding of NAD(P)+-dependent dehydrogenases. The procedure involves preparation of nicotinamide N1-(N-(6-aminohexyl)-acetamide)-agarose and modification of the immobilized nicotinamide by the addition of a ketone or an aldehyde to form an adduct. The nicotinamide, which has no affinity for dehydrogenase, becomes a very specific ligand of dehydrogenase, which binds the ketone or the aldehyde as substrate or inhibitor. In tests, the adduct prepared with immobilized nicotinamide and sodium pyruvate bound specifically to lactate dehydrogenase (EC 1.1.1.27), whereas the adduct prepared with alpha-ketoglutarate bound to glutamate dehydrogenase (EC 1.4.1.3). This technique enables the rapid isolation of a given dehydrogenase.
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PMID:Rapid separation of dehydrogenases by affinity chromatography with new induced specificity phases. 294 22

The time-correlated single photon counting (TCPC) fluorescence technique has been used as a novel approach to investigate ligand-protein interaction, for the case of the binding of the fluorescent coenzyme analogue 1,N6-ethenoNAD (epsilon NAD) to bovine liver glutamate dehydrogenase in the presence of glutarate, a substrate analogue which stabilizes the complex. System calibration was performed using solutions of epsilon ADP and carefully purified epsilon NAD mixed at variable molar ratios (pH 7.0, 0.05 M sodium phosphate buffer, 20 degrees C). The fluorescence lifetimes obtained after deconvolution were 2.4 ns (for epsilon NAD) and 23 ns (for epsilon ADP), in good agreement with literature values obtained under similar conditions. epsilon NAD binds to glutamate dehydrogenase in the presence of 50 mM glutarate, with a fluorescence quantum yield enhancement factor, Q, of about 17-fold, as previously reported (Favilla, R. and Mazzini, A. (1984) Biochim. Biophys. Acta 48-57). For this system, fluorescence lifetime values were obtained after deconvolution as 2.4 ns for free epsilon NAD and 21 ns for bound epsilon NAD. These values did not vary appreciably with enzyme concentration nor with degree of saturation, thus reflecting the existence of only one spectroscopically relevant type of complex. Addition of either GTP or ADP did not affect the lifetime of epsilon NAD bound to the enzyme, but only its affinity, thus allowing calculations of binding strengths. In the case of a simple binding (i.e., in the absence of GTP) the dissociation constant of the complex could be derived from a simple relationship, in which only the ratio between the pre-exponential factors and the parameter gamma, which represents the molar fraction of epsilon NAD molecules free in solution in the open conformation, are to be taken into account. The results are in good agreement with those reported by some of us (reference above) using a steady-state fluorescence technique, which by itself is, however, unable to resolve the number of relevant species present in the system.
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PMID:The binding of 1,N6-ethenoNAD to bovine liver glutamate dehydrogenase: studies using the time-correlated single photon counting fluorescence technique. 348 73

Using fully mechanized analytical equipment, interference by haemolysis in the determination of 26 clinical chemical parameters was determined quantitatively by adding haemolysate to serum. Haemoglobin concentrations up to 6.6 g/l caused essentially no interference in the following determinations: albumin (immuno-nephelometric), alpha-amylase, calcium, chloride, cholesterol, cholinesterase, creatinine, iron, glucose, glutamate dehydrogenase, uric acid, urea, sodium, inorganic phosphate, total protein, transferrin and triglycerides. In the presence of haemoglobin, erroneously high values were found for: lactate dehydrogenase (haemoglobin higher than 0.2 g/l), aspartate aminotransferase, potassium and acid phosphate (haemoglobin higher than 1.5 g/l), creatine kinase (haemoglobin higher than 2.5 g/l) and alanine aminotransferase (haemoglobin higher than 3.4 g/l). Erroneously low values were found for bilirubin (haemoglobin higher than 0.8 g/l), alkaline phosphatase and albumin (by electrophoresis) (haemoglobin higher than 1.5 g/l) and gamma-glutamyltransferase (haemoglobin higher than 3.0 g/l).
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PMID:Haemolysis as an interference factor in clinical chemistry. 371 97

In order to study the intracellular localization of the proteolytic processing steps in the maturation of alpha-glucosidase and cathepsin D in cultured human skin fibroblasts we have used incubation with glycyl-L-phenylalanine-beta-naphthylamide (Gly-Phe-NH-Nap) as described by Jadot et al. [Jadot, M., Colmant, C., Wattiaux-de Coninck, S. & Wattiaux, R. (1984) Biochem. J. 219,965-970] for the specific lysis of lysosomes. When a homogenate of fibroblasts was incubated for 20 min with 0.5 mM Gly-Phe-NH-Nap, a substrate for the lysosomal enzyme cathepsin C, the latency of the lysosomal enzymes alpha-glucosidase and beta-hexosaminidase decreased from 75% to 10% and their sedimentability from 75% to 20-30%. In contrast, treatment with Gly-Phe-NH-Nap had no significant effect on the latency of galactosyltransferase, a marker for the Golgi apparatus, and on the sedimentability of glutamate dehydrogenase and catalase, markers for mitochondria and peroxisomes, respectively. The maturation of alpha-glucosidase and cathepsin D in fibroblasts was studied by pulse-labelling with [35S]methionine, immunoprecipitation, polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate and fluorography. When homogenates of labelled fibroblasts were incubated with Gly-Phe-NH-Nap prior to immunoprecipitation, 70-80% of all proteolytically processed forms of metabolically labelled alpha-glucosidase and cathepsin D was recovered in the supernatant. The earliest proteolytic processing steps in the maturation of alpha-glucosidase and cathepsin D appeared to be coupled to their transport to the lysosomes. Although both enzymes are transported via the mannose-6-phosphate-specific transport system, the velocity with which they arrived in the lysosomes was consistently different. Whereas newly synthesized cathepsin D was found in the lysosomes 1 h after synthesis, alpha-glucosidase was detected only after 2-4 h. When a pulse-chase experiment was carried out in the presence of 10 mM NH4Cl there was a complete inhibition of the transport of cathepsin D and a partial inhibition of that of alpha-glucosidase to the lysosomes. Leupeptin, an inhibitor of lysosomal thiol proteinases, had no effect on the transport of labelled alpha-glucosidase to the lysosomes. However, the early processing steps in which the 110-kDa precursor is converted to the 95-kDa intermediate form of the enzyme were delayed, a transient 105-kDa form was observed and the conversion of the 95-kDa intermediate form to the 76-kDa mature form of the enzyme was completely inhibited.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Biosynthesis and intracellular transport of alpha-glucosidase and cathepsin D in normal and mutant human fibroblasts. 390 6


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