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Query: EC:1.2.1.13 (glyceraldehyde-3-phosphate dehydrogenase)
 6,511 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The spontaneous inactivation of yeast glyceraldehyde-3-phosphate dehydrogenase was found to fit a simple two-state model at pH 8.5 and 25 degrees. The first step is a relatively rapid dissociation of the tetramer to dimers with the equilibrium largely in favor of the tetramer. In the absence of NAD+ the dimer inactivates irreversibly. The apoenzyme is quite stable with a half-life for complete activity loss proportional to the square root of the enzyme concentration. Perturbances of the protein structure (by pH, ionic strength, and specific salts), which have no effect on the tetrameric state of the molecule, result in an alteration of the cooperativity of NAD+ binding, the reactivity of the active-site sulfhydryl group, and the catalytic activity of the enzyme. Covalent modification of two of the four active-site sulfhydryl groups has profound effects on the enzymic activity which are mediated by changes in the subunit interactions. Sedimentation analysis and hybridization studies indicate that the interaction between subunits remains strong after covalent modification. Under normal physiological and equilibrium dialysis conditions the protein is a tetramer. Equilibrium dialysis studies of NAD+ binding to the enzyme at pH 8.5 and 25 degrees reveal a mixed cooperativity pattern. A model consistent with these observations and the observed half-of-the-sites reactivity is that of ligand induced sequential conformational changes which are transferred across strongly interacting subunit domains. Methods for distinguishing negatively cooperative binding patterns from mixtures of denatured enzyme and multiple species are discussed.
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PMID:Subunit interactions in yeast glyceraldehyde-3-phosphate dehydrogenase. 0 55

The mechanisms of glycolytic inhibition in ischemic myocardium were investigated in the isolated, perfused rat heart. Glycolysis was inhibited at the level of glyceraldehyde-3-phosphate dehydrogenase. The major factors that accounted for the glycolytic inhibition in the ischemic heart compared with the anoxic heart appeared to be higher tissue levels of lactate and H+ in the ischemic tissue. Increased extracellular pH inhibited glycolysis in anoxic and hypoxic hearts much more readily than it did in aerobic hearts. However, maintenance of both extracellular and intracellular pH caused only a modest acceleration of glycolysis in ischemic hearts. Accumulation of tissue lactate and inhibition of glycolysis were directly proportional to the reduction in coronary bloow flow in both anoxic and ischemic hearts. At intracellular lactate concentrations between 15 and 20 mM, glycolysis was inhibited under both conditions. Addition of either 10, 20, or 40 mM lactate to the perfusate inhibited glycolysis in aerobic, anoxic, and ischemic hearts. The effect of lactate did not appear to be mediated through changes in intracellular pH. It is concluded that accumulation of lactate represents a major factor in the inhibition of glycolysis that develops in ischemic hearts.
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PMID:Mechanisms of glycolytic inhibition in ischemic rat hearts. 0 Jan 57

NADH and NADPH-ferredoxin oxidoreductases have been studied in Clostridium acetobutylicum, Cl. tyrobutyricum and Cl. pasteurianum. The study of the distribution and regulation of these enzymatic activities in well-defined culture conditions, reveals that the essential function of NADPH-ferredoxin oxidoreductase is to produce NADPH, while NADH-ferredoxin oxidoreductase can, depending on cellular conditions, produce or oxidize NADH. When these Clostridia use glycolysis, regulation of the NADH-ferredoxin oxidoreductase by acetyl-CoA (obligatory activator of NADH-ferroxin reductase activity) and by NADH (competitive inhibitor of ferredoxin-NAD+ reductase activity) allow the enzymes to function correlatively with glyceraldehyde-3-phosphate dehydrogenase and thus control the levels of NAD+ and NADH in the cell. In Cl. tyrobutyricum and Cl. pasteurianum, the ferredoxin-NADP+ reductase activities are regulated by NAD+ and NADH in accordance with the intracellular concentrations of these coenzymes. In Cl. tyrobutyricum growing on pyruvate/acetate, NADH and NADPH-ferredoxin reductase activities cannot be detected; only the ferredoxin-NAD+ and ferredoxin-NADP+ reductase activities are found. In this Clostridium, regulation of the ferredoxin-NADP+ reductase activity is the same whether it is grown on glucose or pyruvate. Contrary to this, the ferredoxin-NAD+ reductase activity undergoes a drastic change, since NADH no longer controls the enzymatic activity. In this case regulation is no longer necessary, since glyceraldehyde-3-phosphate dehydrogenase does not function.
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PMID:Regulation of the NADH and NADPH-ferredoxin oxidoreductases in clostridia of the butyric group. 0 18

Two enzymes with glyceraldehyde-3-phosphate dehydrogenase activity have been purified from heterotrophically grown Scenedesmus obliquus by ion-exchange chromatography and gel filtration. The D-enzyme has a molecular weight of 550000 and a VNADH: VNADPH ratio of 16 whereas the T-enzyme has a molecular weight of 140000 and a VNADH:VNADPH ratio of 0.15. The two enzymes, however, are very similar with regard to their Michaelis constants for the reduced pyridine nucleotides, pH optimum, subunit size and ultraviolet absorption.
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PMID:Algal glyceraldehyde-3-phosphate dehydrogenase. Pyridine-nucleotide requirements of two enzymes purified from Scenedesmus obliquus. 0 11

Incubation of HeLa cells with [32P]orthophosphate results in more rapid labeling of the gamma-phosphorus of ATP than of the intracellular pool of orthophosphate. The specific radioactivity of ATP equals that of extracellular orthophosphate after 2 h of incubation. A similar pattern of labeling is seen with human erythrocytes when incubated at physiological concentrations of orthophosphate (2 mM) and pH 7.4-7.8. At lower pH, 6.8-7.2, the rate of orthophosphate uptake increases and exceeds the rate of labeling of ATP. These data are explained by the existence of a primary system for ATP uptake which involves the mediation of membrane-bound glyceraldehyde-3-phosphate dehydrogenase. Phosphate first enters the cell as 1,3-diphosphoglyceric acid, is then transferred to ATP, and then enters the intracellular orthophosphate pool. At lower pH monovalent orthophosphate also enters the erythrocyte by a process not involving glyceraldehyde-3-phosphate dehydrogenase.
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PMID:Mode of orthophosphate uptake and ATP labeling by mammalian cells. 0 42

Chloroplast NADP-linked glyceraldehyde-3-phosphate dehydrogenase was resolved into three forms that differed in molecular weight: (a) larger than or equal to 1.5 million; (b) 600,000; and (c) less than or equal to 100,000. After preincubation with an effector (ATP, NADPH, or Pi) the activity of forms a and c was unaffected, whereas the activity of b, the regulatory form, was increased 10-fold. Activation was accompanied by the exposure of previously hidden sulfhydryl groups. The rate of activation was slower than the rate of catalysis and resulted in a lag phase during the measurement of activity when the enzyme was preincubated in the absence of an effector. The addition of one of several compounds as a second effector (at a concentration which itself was nonactivating) in the presence of a first effector enhanced activation by lowering the concentration of the first effector required for half-maximal activation (Pi constant/ATP or NADPH varied; ATP or NADPH constant/Pi varied). Other combinations of effectors caused little change in activity (ATP constant/NADPH varied; NADPH constant/ATP varied). Glyceraldehyde 3-phosphate added as a second effector induced contrasting changes: an increase in the ATP-mediated activation and a decrease in the NADPH-mediated activation. The results are consistent with the view that the products of the photochemical reactions of chloroplasts, ATP, and NADPH, in conjunction with other metabolites, regulate the activity of glyceraldehyde-3-phosphate dehydrogenase in the photosynthetic assimilation of CO2.
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PMID:Studies on the regulation of chloroplast NADP-linked glyceraldehyde-3-phosphate dehydrogenase. 1 Feb 97

The binding of nicotinamide adenine dinucleotide (NAD+) to yeast glyceraldehyde-3-phosphate dehydrogenase (GPDH) has been studied at pH 6.5 and 8.5, at 5,25, and 40 degrees C, by calorimetry, fluorometry, spectrophotometry, equilibrium dialysis, and flow dialysis. As reported earlier for pH 7.3 (Velick S.F., Baggott, J.P., and Sturtevant, J.M. (1971), Biochemistry 10, 779), the binding is accompanied by enthalpy changes which become rapidly more negative as the temperature increases, with delta Cp = -500 to -750 cal deg-1 (mole of NAD+ bound)-1, and by entropy changes which also, as required by the large negative delta Cp, become rapidly more negative with increasing temperature. The binding data at pH 6.5 can be fitted on the basis of either four identical noninteracting sites, or of four sites showing a small degree of negative cooperativity. The data at pH 8.5, particularly at 40 degrees C, require the introduction of positive cooperativity, as was previously shown by Kirschner et al. (Kirschner, K., Eigen, M., Bittman, R., and Voigt, B. (1966), Proc. Natl. Acad. Sci. U.S.A. 56, 1661), and can be equally well fitted on the basis of a sequential model (Adair, G.S. (1925), J. Biol. Chem. 63, 529) or a concerted model (Monod, J., Wyman, J., and Changeux, J.P. (1965), J. Mol. Biol. 12, 88). It is proposed that the observed thermodynamic changes are largely the result of a hydrophobic effect due to a decrease in the exposure of nonpolar groups to the solvent, and of a tightening of the protein structure when the coenzyme is bound with concomitant decrease in the number of easily excitable internal degrees of freedom.
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PMID:Energetics of the cooperative and noncooperative binding of nicotinamide adenine dinucleotide to yeast glyceraldehyde-3-phosphate dehydrogenase at pH 6.5 and pH 8.5. Equilibrium and calorimetric analysis over a range of temperature. 1 17

NADH-dependent glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.--) of the photosynthetic alga Scenedesmus obliquus is converted to an NADPH specific form by incubation with dithiothreitol. The change in nucleotide specificity is accompanied by a reduction in the molecular weight of the enzyme from 550 000 to 140 000. Prolonged incubation with dithiothreitol results in the further dissociation of the enzyme to an inactive 70 000 dalton species. The 140 000 dalton, NADPH-specific enzyme is stabilized against dissociation and inactivation by the presence of NAD(H) or NADP(H). Optimum stimulation of NADPH-dependent glyceraldehyde-3-phosphate dehydrogenase activity is achieved on incubation of the NADH-specific enzyme with dithiothreitol and NADPH, or dithiothreitol and a 1,3-diphosphoglycerate generating system. The relevance of these observations to in vivo light-induced changes in the nucleotide specificity of the enzyme is discussed.
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PMID:Glyceraldehyde-3-phosphate dehydrogenase of Scenedesmus obliquus. Effects of dithiothreitol and nucleotide on coenzyme specificity. 1 3

The formation of ternary Cu-enzyme-coenzyme complex from cupric ion and D-glyceraldehyde-3-phosphate dehydrogenase holoenzyme results in similar spectral changes as the formation of binary Cu-apoenzyme complex, which indicates that the complex bonds between cupric ion and the holoenzyme, and cupric ion and the apoenzyme are similar. Spectrophotometric titration, chemical modification experiments and inhibition studies with cupric ion gave evidence that cupric ion is selectively bound on Cys-149 residue also in the Cu-GAPD-NAD complex. The charge transfer interaction between the coenzyme and Cu-GAPD, i.e. the difference spectrum of the combination of NAD with Cu-GAPD complex, is different from that of the enzyme-coenzyme complex in the absence of cupric ion. The shape of this "modified enzyme-coenzyme charge transfer spectrum" is influenced by various anions. The difference absorption does not depend on the pH in the range of 5.5 to 9. This indicates that the bound cupric ion abolishes the effect of deprotonation of a functional group in the protein on the charge transfer interaction. It is suggested that this functional group is a histidine imidazole, which activates the Cys-149 thiol group in the native enzyme and binds the metal ion in the cupric complex in a Cys-Cu-His chelate structure.
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PMID:Complex of D-glyceraldehyde-3-phosphate dehydrogenase with Cu2+ ion. The properties of ternary Cu-enzyme-coenzyme complex. 1 25

Procedure for isolation of electrophoretically homogeneous, crystalline glyceraldehyde-3-phosphate dehydrogenase from lamprey muscles is described. Amino acid composition of the enzyme was investigated and compared with that of the same dehydrogenase from other sources. With respect to its secondary structure and kinetic parameters, the lamprey enzyme does not significantly differ from those of other animals. Activation energy for the lamprey enzyme is lower than for the same enzyme from endothermic animasl. "Temperature modulation" of Michaelis constant described in the literature, was not confirmed for the lamprey enzyme in the range of physiological pH values.
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PMID:[Isolation and several properties of the glyceraldehyde-3-phosphate dehydrogenase from the muscles of the lamprey Lampetra fluviatillis]. 1 52


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