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

Random collisions between macromolecules lead to dynamic associations (lengthy encounters) that in principle could affect their conformation and, in the case of enzymes, their binding and catalytic properties. Exploiting the unique sensitivity of the phosphorescence lifetime, tau, of Trp to the internal flexibility of globular proteins we probed the perturbations induced in the structure of the coenzyme-binding domain of alcohol dehydrogenase (LADH) and glyceraldehyde-3-phosphate dehydrogenase (GraPDH) by the presence in solution of other dehydrogenases and of functionally unrelated proteins. With Trp314 of LADH, the results emphasize that while tau is not affected by the concentration of LADH itself, the addition of micromolar quantities of other proteins causes a distinct reduction in it. From the linear increase of 1/tau with protein concentration one obtains values for the apparent second-order Stern-Volmer rate constant that range between 2-200 x 10(3) M-1 s-1, decreasing 2-3-fold when ternary complexes of LADH with NADH or NAD+ and inhibitors are involved. Similar effects were observed with Trp310 of GraPDH except that with sorbitol dehydrogenase as perturbant the increase of 1/tau is hyperbolic and governed by an apparent dissociation constant of about 1 microM. Finally, glycerol-3-phosphate dehydrogenase, the strongest perturber of both LADH and GraPDH, has either no effect on lactic dehydrogenase from pig heart or induces a moderate lengthening of the triplet lifetime of the rabbit muscle enzyme. Because Stern-Volmer behavior is typical also of diffusion-mediated quenching reactions, a parallel investigation with cysteine, cystine and N-acetyl-tryptophanamide demonstrated that among potential, protein-associated, quenching moieties namely, -SH, -S-S- and indole groups, only the latter has rate constants approaching the magnitude of protein perturbants. Since considerable evidence rules out the predominance of such quenching reactions, these findings confirm a subtle form of communication between protein molecules in solution. The lack of specificity and the similar effects between dehydrogenases with right and wrong stereospecificity for direct coenzyme transfer suggests that the perturbations monitored are unrelated to this function.
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PMID:Conformational changes in proteins induced by dynamic associations. A tryptophan phosphorescence study. 816 51

Incubation of glyceraldehyde-3-phosphate dehydrogenase (GAPD) with sodium nitroprusside (SNP) decreased its activity in concentration- and time-dependent fashion in the presence of a thiol compound, with DTT being more effective than GSH. Both forward and backward reactions were effected. Coinciding with this, HgCl2-sensitive labelling of the protein by [32P]NAD+ also increased, indicating the stimulation of ADP-ribosylation. Treatment with SNP of GAPD samples from rabbit muscle, sheep brain and yeast inactivated the dehydrogenase activity of the three, but only the mammalian proteins showed ADP-ribosylation activity. The SNP-modified protein of rabbit muscle GAPD, freed from the reagent by Sephadex filtration showed a concentration-dependent restoration of the dehydrogenase activity on preincubation with DTT and GSH. Such thiol-treated preparations also gave increased ADP-ribosylation activity with DTT, and to a lesser extent with GSH. The SNP-modified protein was unable to catalyze this activity with the native yeast enzyme and native and heat-inactivated muscle enzyme. It was possible to generate the ADP-ribosylation activity in muscle GAPD, by an NO-independent mechanism, on dialysis in Tris buffer under aerobic conditions, and on incubating with NADPH, but not NADH, in muscle and brain, but not yeast, enzymes. The results suggest that the inverse relationship of the dehydrogenase and ADP-ribosylation activities is coincidental but not correlated.
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PMID:Inverse relationship of the dehydrogenase and ADP-ribosylation activities in sodium-nitroprusside-treated glyceraldehyde-3-phosphate dehydrogenase is coincidental. 821 90

Sulfenic acids (R-SOH) result from the stoichiometric oxidations of thiols with mild oxidants such as H2O2; in solution, however, these derivatives accumulate only transiently due to rapid self-condensation reactions, further oxidations to the sulfinic and/or sulfonic acids, and reactions with nucleophiles such as R-SH. In contrast, oxidations of cysteinyl side chains in proteins, where disulfide bond formation can be prevented and where the reactivity of the nascent cysteine-sulfenic acid (Cys-SOH) can be controlled, have previously been shown to yield stable active-site Cys-SOH derivatives of papain and glyceraldehyde-3-phosphate dehydrogenase. More recently, however, functional Cys-SOH residues have been identified in the native oxidized forms of the FAD-containing NADH peroxidase and NADH oxidase from Streptococcus faecalis; these two proteins constitute a new class within the flavoprotein disulfide reductase family. In addition, Cys-SOH derivatives have been suggested to play important roles in redox regulation of the DNA-binding activities of transcription factors such as Fos and Jun, OxyR, and bovine papillomavirus type 1 E2 protein. Structural inferences for the stabilization of protein-sulfenic acids, drawn from the refined 2.16-A structure of the streptococcal NADH peroxidase, provide a molecular basis for understanding the proposed redox functions of these novel cofactors in both enzyme catalysis and transcriptional regulation.
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PMID:Protein-sulfenic acid stabilization and function in enzyme catalysis and gene regulation. 826 33

The nucleotide sequence of trkA, a gene encoding a surface component of the constitutive K(+)-uptake systems TrkG and TrkH from Escherichia coli, was determined. The structure of the TrkA protein deduced from the nucleotide sequence accords with the view that TrkA is peripherally bound to the inner side of the cytoplasmic membrane. Analysis by a dot matrix revealed that TrkA is composed of similar halves. The N-terminal part of each TrkA half (residues 1-130 and 234-355, respectively) is similar to the complete NAD(+)-binding domain of NAD(+)-dependent dehydrogenases. The C-terminal part of each TrkA half (residues 131-233 and 357-458, respectively) aligns with the first 100 residues of the catalytic domain of glyceraldehyde-3-phosphate dehydrogenase. Strong u.v. illumination at 252 nm led to cross-linking of NAD+ or NADH, but not of ATP to the isolated TrkA protein.
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PMID:NAD+ binding to the Escherichia coli K(+)-uptake protein TrkA and sequence similarity between TrkA and domains of a family of dehydrogenases suggest a role for NAD+ in bacterial transport. 841

A wide variety of cinnamic acid derivatives are inhibitors of the low Km mitochondrial aldehyde dehydrogenase. Two of the most potent inhibitors are alpha-cyano-3,4-dihydroxythiocinnamamide (Ki0.6 microM) and alpha-cyano-3,4,5-trihydroxycinnamonitrile (Ki2.6 microM). With propionaldehyde as substrate the inhibition by these compounds was competitive with respect to NAD+. alpha-Fluorocinnamate was a much less effective inhibitor of the enzyme, with mixed behaviour towards NAD+, but with a major competitive component. These cinnamic acid derivatives were ineffective as inhibitors of the aldehyde dehydrogenase-catalysed hydrolysis of p-nitrophenyl acetate, but inhibited the ability of NAD+ and NADH to activate this activity. Inhibition of the stimulation of esterase activity was competitive with respect to NAD+ and NADH, and the derived Ki values were the same as for inhibition of dehydrogenase activity. NAD+, but not acetaldehyde, could elute the low Km aldehyde dehydrogenase from alpha-cyanocinnamate-Sepharose, to which the enzyme binds specifically (Poole RC and Halestrap AP, Biochem J 259: 105-110, 1989). The cinnamic acid derivatives have little effect on lactate dehydrogenase, glyceraldehyde-3-phosphate dehydrogenase or a high Km aldehyde dehydrogenase present in rat liver mitochondria. It is concluded that some cinnamic acid derivatives are potent inhibitors of the low Km aldehyde dehydrogenase, by competing with NAD+/NADH for binding to the enzyme. They are much less effective as inhibitors of other NAD(+)-dependent dehydrogenases.
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PMID:Derivatives of cinnamic acid interact with the nucleotide binding site of mitochondrial aldehyde dehydrogenase. Effects on the dehydrogenase reaction and stimulation of esterase activity by nucleotides. 848 2

To evaluate the relationship of inositol 1,4,5-trisphosphate (IP3) receptor-mediated signal transduction and cellular energy dynamics, we have characterized effects of nucleotides on IP3 receptor (IP3R)-mediated calcium (Ca2+) flux in purified IP3 receptors reconstituted in lipid vesicles (IP3RV) and examined hypoxia-induced augmentation of intracellular Ca2+ in intact cells. Reduced nicotinamide adenine dinucleotide (NADH) increases IP3-mediated Ca2+ flux in IP3RV. This effect is highly specific for NADH. Hypoxia elicited by brief exposure of nerve growth factor-differentiated PC12 cells or cerebellar Purkinje cells to cyanide elicits rapid increased in internal [Ca2+], which derives from IP3-sensitive stores. Blockade of this effect by 2-deoxyglucose and inhibition of glyceraldehyde-3-phosphate dehydrogenase implicates enhanced glycolytic production of NADH in the Ca2+ stimulation. Internal [Ca2+] is markedly and specifically increased by direct intracellular injection of NADH, and this effect is blocked by heparin, further implicating IP3R stores. These findings indicate that direct regulation of IP3R by NADH is responsible for elevated cytoplasmic [Ca2+] occurring in the earliest phase of hypoxia. This link of IP3R activity with cellular energy dynamics may be relevant to both hypoxic damage and metabolic regulation of IP3 signaling processes.
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PMID:Reduced nicotinamide adenine dinucleotide-selective stimulation of inositol 1,4,5-trisphosphate receptors mediates hypoxic mobilization of calcium. 860 44

Nitric oxide (NO)-related activity has been associated with an NAD+-dependent modification of the glycolytic enzyme, glyceraldehyde-3-phosphate dehydrogenase (GAPDH). However, the mechanism by which NO effects covalent attachment of nucleotide and its role in regulation of enzyme activity are controversial. Recent studies have shown that S-nitrosylation of GAPDH (Cys149) initiates subsequent modification by the pyridinium cofactor. Here we show that NADH rather than NAD+ is the preferred substrate. Transnitrosation from active site S-nitrosothiol to the reduced nicotinamide ring system appears to facilitate protein thiolate attack on the enzyme-bound cofactor. This results in attachment of the intact NADH molecule. Moreover, we find that S-nitrosylation of GAPDH is responsible for reversible enzyme inhibition, whereas attachment of NADH accounts for irreversible enzyme inactivation. S-Nitrosylation may serve to protect GAPDH from oxidant inactivation in settings of cytokine overproduction and to regulate glycolysis. NADH attachment is more likely to be a pathophysiological event associated with inhibition of gluconeogenesis.
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PMID:Posttranslational modification of glyceraldehyde-3-phosphate dehydrogenase by S-nitrosylation and subsequent NADH attachment. 862 64

The subcellular localization of l-lactate dehydrogenase (LDH) in rat hepatocytes has been studied by analytical subcellular fractionation combined with the immunodetection of LDH in isolated subcellular fractions and liver sections by immunoblotting and immunoelectron microscopy. The results clearly demonstrate the presence of LDH in the matrix of peroxisomes in addition to the cytosol. Both cytosolic and peroxisomal LDH subunits have the same molecular mass (35.0 kDa) and show comparable cross-reactivity with an anti-cytosolic LDH antibody. As revealed by activity staining or immunoblotting after isoelectric focussing, both intracellular compartments contain the same liver-specific LDH-isoforms (LDH-A4 > LDH-A3B) with the peroxisomes comprising relatively more LDH-A3B than the cytosol. Selective KCl extraction as well as resistance to proteinase K and immunoelectron microscopy revealed that at least 80% of the LDH activity measured in highly purified peroxisomal fractions is due to LDH as a bona fide peroxisomal matrix enzyme. In combination with the data of cell fractionation, this implies that at least 0.5% of the total LDH activity in hepatocytes is present in peroxisomes. Since no other enzymes of the glycolytic pathway (such as phosphoglucomutase, phosphoglucoisomerase, and glyceraldehyde-3-phosphate dehydrogenase) were found in highly purified peroxisomal fractions, it does not seem that LDH in peroxisomes participates in glycolysis. Instead, the marked elevation of LDH in peroxisomes of rats treated with the hypolipidemic drug bezafibrate, concomitantly to the induction of the peroxisomal beta-oxidation enzymes, strongly suggests that intraperoxisomal LDH may be involved in the reoxidation of NADH generated by the beta-oxidation pathway. The interaction of LDH and the peroxisomal palmitoyl-CoA beta-oxidation system could be verified in a modified beta-oxidation assay by adding increasing amounts of pyruvate to the standard assay mixture and recording the change of NADH production rates. A dose-dependent decrease of NADH produced was simulated with the lowest NADH value found at maximal LDH activity. The addition of oxamic acid, a specific inhibitor of LDH, to the system or inhibition of LDH by high pyruvate levels (up to 20 mm) restored the NADH values to control levels. A direct effect of pyruvate on palmitoyl-CoA oxidase and enoyl-CoA hydratase was excluded by measuring those enzymes individually in separate assays. An LDH-based shuttle across the peroxisomal membrane should provide an efficient system to regulate intraperoxisomal NAD+/NADH levels and maintain the flux of fatty acids through the peroxisomal beta-oxidation spiral.
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PMID:L-lactate dehydrogenase A4- and A3B isoforms are bona fide peroxisomal enzymes in rat liver. Evidence for involvement in intraperoxisomal NADH reoxidation. 863 3

NADP-dependent glyceraldehyde-3-phosphate dehydrogenase (GAPDH) has been purified to electrophoretic homogeneity from Synechococcus PCC 7942 cells. The native enzyme had a molecular mass of 160 kDa and consisted of four subunits with a molecular mass of 41 kDa. The activity was 6-fold higher with NADPH than with NADH; the apparent Km values for NADPH and NADH were 62 +/- 4.5 and 420 +/- 10.5 microM respectively. The gene encoding NADP-dependent GAPDH was cloned from the chromosomal DNA of Synechococcus 7942. A 1140 bp open reading frame, encoding an enzyme of 380 amino acid residues (approx.molecular mass of 41.3 kDa) was observed. The deduced amino acid sequence of the gene had a greater sequence similarity to the NADP-dependent and chloroplastic form than to the NAD-dependent and cytosolic form. The Synechococcus 7942 enzyme lacked one of the cysteines involved in the light-dependent regulation of the chloroplast enzymes of higher plants. The recombinant enzyme expressed in Escherichia coli as well as the native enzyme purified from Synechococcus 7942 cells were resistant to 1 mM H2O2.
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PMID:Enzymic and molecular characterization of NADP-dependent glyceraldehyde-3-phosphate dehydrogenase from Synechococcus PCC 7942: resistance of the enzyme to hydrogen peroxide. 868 18

The effects of troglitazone and pioglitazone on glucose and fatty acid metabolism were studied in hepatocytes isolated from 24-h-starved rats. These thiazolidinediones inhibited long-chain fatty acid (oleate) oxidation and produced a very oxidized mitochondrial redox state. By contrast, thiazolidinediones did not affect the rate of medium-chain fatty acid (octanoate) oxidation or the activity of mitochondrial carnitine palmitoyltransferase (CPT) I. Thiazolidinediones inhibited selectively triglyceride synthesis but not phospholipid synthesis. The combined inhibition of oleate oxidation and esterification by troglitazone was due to a noncompetitive inhibition of mitochondrial and microsomal long-chain acyl-CoA synthetase (ACS) activities. It was suggested that troglitazone must be metabolized into its sulfo-conjugate derivative in liver cells to inhibit mitochondrial and microsomal ACS activities. Thiazolidinediones inhibited glucose production from lactate/pyruvate or from alanine. Analysis of gluconeogenic metabolite concentrations suggested that troglitazone would inhibit gluconeogenesis at the level of pyruvate carboxylase and glyceraldehyde-3-phosphate dehydrogenase reactions. It was concluded that 1) at a similar concentration, troglitazone was more efficient than pioglitazone to inhibit fatty acid metabolism and gluconeogenesis and 2) the inhibition of gluconeogenesis by troglitazone could be the result of the inhibition of long-chain fatty acid oxidation (decrease in acetyl-CoA, NADH-to-NAD+, and ATP-to-ADP ratios).
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PMID:Troglitazone inhibits fatty acid oxidation and esterification, and gluconeogenesis in isolated hepatocytes from starved rats. 886 61


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