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)

Nitric oxide was recently demonstrated to stimulate ADP-ribosylation of glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Our studies on the effect of glyceraldehyde-3-phosphate (GA3P), the natural substrate of dehydrogenase activity of GAPDH, indicated GA3P to be another very potent activator of ADP-ribosylation of the enzyme. GA3P was able to activate ADP-ribosylation only in the presence of DTT. The action of GA3P was associated with inhibition of GAPDH dehydrogenase activity. Ka for GA3P was at least 50-fold lower and maximal activation was somewhat higher than these values for other aldehydes that were also able to enhance GAPDH ADP-ribosylation in the presence of DTT. ADP-ribosylation was blocked by carboxamidomethylation of the essential cysteine SH-group. The bond between the prelabeled protein and ADP-ribose was resistant to hydrolysis with hydroxylamine and HgCl2, suggesting that a lysine epsilon-amino group is the target for ADP-ribosylation.
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PMID:Glyceraldehyde-3-phosphate activates auto-ADP-ribosylation of glyceraldehyde-3-phosphate dehydrogenase. 850 56

D-Glyceraldehyde irreversibly inhibited rat liver glucokinase in a concentration-dependent manner. The inactivation of glucokinase by glyceraldehyde was blocked by the presence of its substrates such as glucose and mannose. Glucokinase was highly sensitive to glyceraldehyde compared with some other glycolytic enzymes (from animal tissues) including hexokinase, glucose-6-phosphate isomerase, 6-phosphofructokinase, glyceraldehyde-3-phosphate dehydrogenase, and pyruvate kinase. The amino acid analysis of untreated and glyceraldehyde-treated glucokinase suggested that glyceraldehyde-induced inactivation of glucokinase is caused by glycation of Lys residues of the enzyme by the triose. Treatment of pancreatic islets with 6 mM glyceraldehyde for 1 h at 37 degrees C caused both inactivation of glucokinase and inhibition of glucose-induced insulin secretion. Another glucose-phosphorylating enzyme (hexokinase) in pancreatic islets, however, was little affected by glyceraldehyde. In addition, glyceraldehyde did not affect the insulin secretory responses of islets to nonglucose secretagogues such as glyceraldehyde and Leu. When pancreatic islets were cultured with a lower concentration (1 mM) of glyceraldehyde for a longer time (17 h) in the presence of 10 mM glucose to mimic the in vivo conditions, both glucokinase activity and glucose-induced insulin secretion were again decreased. This study demonstrates that glucose-induced insulin secretion is impaired by glyceraldehyde through the inactivation of glucokinase. The implication of this finding in the pathophysiology of type II diabetes is discussed.
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PMID:Inhibition of glucose-induced insulin secretion through inactivation of glucokinase by glyceraldehyde. 851 67

The role of the protein tyrosine kinase pp60c-src in the expression of prostaglandin G/H synthase (PGHS), the key enzyme of prostaglandin synthesis, was investigated in rat renal mesangial cells. Transfection of mesangial cells with the proto-oncogene c-src resulted in nontransformed cells with constitutively enhanced pp60c-src kinase activity. As a control, mesangial cells were transfected with inactive pp60c-src, mutated in position 295 (lysine replaced by methionine). Expression of the constitutive isoform PGHS-1 was enhanced in cells overexpressing wild-type c-src compared to cells transfected with the kinase negative c-src mutant. Levels of other constitutively expressed proteins such as GAPDH and beta-actin were also enhanced. PGHS-2 was barely detectable in resting cells but was inducible by PDGF-AB, PDGF-BB, serotonin, FCS, and calcium ionophore A23187. Induction was diminished in pp60c-src kinase-overexpressing cells, independent of the stimulus used, suggesting interference at a late step in the signaling cascade. No induction of PGHS-2 mRNA was observed in mesangial cells transformed by the oncogene v-src. An increase in intracellular calcium levels is an early step in signal transduction by PDGF and serotonin in mesangial cells. c-src kinase overexpression reduced PDGF- and serotonin-mediated changes in Ca2+ signaling, indicating multiple targets of pp60c-src action. Overexpression of pp60c-src in mesangial cells thus affected basal protein expression, reflected by the enhanced PGHS-1 mRNA and protein expression. With regard to induction of PGHS-2, overexpression of pp60c-src reduced induction by stimuli coupled to different types of signaling pathways.
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PMID:Modulation of prostaglandin G/H synthase expression in mesangial cells transfected by pp60c-src proto-oncogene. 859 18

We previously identified DNA sequences involved in the function of the complex promoter of the streptokinase gene from Streptococcus equisimilis H46A, a human serogroup C strain known to express this gene at a high level. As a prerequisite to understanding possible mechanisms that control the balance between the plasminogen activating and plasmin(ogen) binding capacities of H46A, we describe here its gapC gene encoding glyceraldehyde-3-phosphate dehydrogenase (GraP-DH, EC 1.2.1.12), a glycolytic enzyme apparently transported to the cell surface where it functions as a plasmin(ogen).binding protein. The gapC gene was cloned and sequenced and found to code for a 336-amino-acid polypeptide (approximately 35.9 kDa) exhibiting 94.9% sequence identity to the Plr protein from Streptococcus pyogenes shown by others to be capable of plasmin binding [Lottenberg, R., Broder, C. C., Boyle, M. D., Kain, S. J., Schroeder, B. L. & Curtiss, R. III (1992) J. Bacteriol. 174, 5204-5210]. To study the properties of the GapC protein, its gene was inducibly overexpressed in Escherichia coli from QIAexpress expression plasmids to yield the authentic GapC or (His)6GapC carrying a hexahistidyl N-terminus to permit affinity purification. Both proteins were functionally active, exhibiting specific GraP-DH activities of about 80 kat/mol (approximately 130 U/mg) after purification. Their binding parameters [association (ka) and dissociation (kd) rate constants, and equlibrium dissociation constants (Kd = kd/ka)] for the interaction with human Gluplasminogen and plasmin were determined by real-time biospecific interaction analysis using the Pharmacia BIAcore instrument. For comparative purposes, the commercial GraP-DH from Bacillus stearothermophilus (BstGraP-DH), a nonpathogenic organism, was included in these experiments. The Kd values for binding of plasminogen to GapC, (His)6GapC and BstGraP-DH were 220 nM, 260 nM and 520 nM, respectively, as compared to 25 nM, 17 nM and 98 nM, respectively, for the binding to plasmin. These data show that both the zymogen and active enzyme possess low-affinity binding sites for the gapC gene product and that the hexahistidyl terminus does not affect its function. Prior limited treatment with plasmin enhanced the subsequent plasminogen binding capacity of all three GraP-DHs, presumably by the exposure of new C-terminal lysine residues for binding to the zymogen.
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PMID:Cloning, sequencing and functional overexpression of the Streptococcus equisimilis H46A gapC gene encoding a glyceraldehyde-3-phosphate dehydrogenase that also functions as a plasmin(ogen)-binding protein. Purification and biochemical characterization of the protein. 870 17

A mutant phosphoribulokinase has been isolated from the 12-2B mutant of Chlamydomonas reinhardtii. In this mutant, Arg64 has been replaced by Cys. The enzyme, which may exist in the dimeric and tetrameric states, is almost devoid of activity. Neither of these enzymes is able to form a complex with glyceraldehyde-3-phosphate dehydrogenase. The phosphoribulokinase gene has been expressed in Escherichia coli. The resulting recombinant protein, after isolation and purification, is apparently identical to the native enzyme extracted from the chloroplast. Three mutants have been generated by site directed mutagenesis. Arg64 has been replaced by Ala, Lys or Glu. With the exception of the latter, the two other mutants, [A64]phosphoribulokinase and [K64]phosphoribulokinase, are active when they are reduced, and nearly totally inactive in their oxidized state. Their activity, however, is decreased relative to that of the native, or to that of the wild-type recombinant phosphoribulokinase. Both the catalytic constant and the apparent affinity of ribulose 5-phosphate are decreased relative to the corresponding values obtained for the wild-type, the native or the recombinant enzyme. Whereas the [A64]phosphoribulokinase is unable to form a complex with glyceraldehyde-3-phosphate dehydrogenase, [K64]phosphoribulokinase does, but the stability of the resulting complex is much decreased relative to that of the wild-type complex. The oxidized mutant [K64]phosphoribulokinase becomes active in the presence of glyceraldehyde-3-phosphate dehydrogenase but this activity is smaller than that of the corresponding wild-type enzyme. Taken together, these results show that Arg64 plays a major role in the association of the two enzymes and in the information transfer that takes place between glyceraldehyde-3-phosphate dehydrogenase and phosphoribulokinase. As this residue also appears to be important for catalytic activity, it may be tempting to consider that it stabilizes a conformation that is required for both the catalytic activity and the formation of the bienzyme complex.
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PMID:Information transfer in multienzyme complexes--2. The role of Arg64 of Chlamydomonas reinhardtii phosphoribulokinase in the information transfer between glyceraldehyde-3-phosphate dehydrogenase and phosphoribulokinase. 942 76

The aim of the current article is to overview the recent developments in the field of hemorrhagic shock research, as it relates to the roles of nitric oxide (NO) in the pathogenesis of this condition. The first part of the review focuses on the roles of peroxynitrite, a reactive oxidant produced from the reaction of NO and superoxide. The second part of the review deals with the novel findings related to the recently identified regulatory roles of the inducible isoform of nitric oxide synthase (iNOS) in the expression of pro-inflammatory mediators in hemorrhagic shock. (1) The role of peroxynitrite: Immunohistochemical and biochemical evidence demonstrate the production of peroxynitrite in hemorrhagic shock. Peroxynitrite can initiate a wide range of toxic oxidative reactions. These include initiation of tyrosine nitration, lipid peroxidation, direct inhibition of mitochondrial respiratory chain enzymes, inactivation of glyceraldehyde-3-phosphate dehydrogenase, inhibition of membrane sodium/potassium ATP-ase activity, inactivation of membrane sodium channels, and other oxidative modifications of proteins. All these toxicities are likely to play a role in the pathophysiology of hemorrhagic shock. A combined anti-inflammatory agent, mercaptoethylguanidine, which selectively inhibits iNOS and scavenges peroxynitrite, prevents the delayed vascular decompensation and the cellular energetic failure associated with late hemorrhagic shock. Peroxynitrite is a potent trigger of DNA single strand breakage, with subsequent activation of the nuclear enzyme poly (ADP ribose) synthetase (PARS), leading to eventual severe energy depletion of the cells, and necrotic-type cell death. Pharmacological inhibition of PARS, with 3-aminobenzamide or 5-iodo-6-amino-1,2-benzopyrone, improves hemodynamic status and prolongs survival time in rodent and porcine models of severe hemorrhagic shock. (2) Novel signaling roles of induced NO in hemorrhagic shock. Although the severity and duration of shock may dictate the timing and extent of iNOS expression, it is now evident that the up-regulation of iNOS can take place during sustained shock. Accumulated data indicate that iNOS expressed during shock contributes to vascular decompensation, as classically described by Wiggers. In addition, the presence of even low levels of iNOS at the time of resuscitation enhances the inflammatory response that follows the reperfusion state. Pharmacological inhibition of iNOS with N6-(iminoethyl)-L-lysine or genetic inactivation of iNOS (iNOS knockout mice) attenuates the activation of the transcription factors nuclear factor kappa B (NFkappaB) and Signal Transducer and Activator of Transcription 3 (STAT3), and ameliorates the increases in interleukin-6 and G-CSF messenger RNA levels in the lungs and liver. Inhibition of iNOS results in a marked reduction of lung and liver injury produced by hemorrhagic shock. Thus, induced nitric oxide, in addition to being a "final common mediator" of hemorrhagic shock, is essential for the up-regulation of the inflammatory response in resuscitated hemorrhagic shock. Furthermore, a picture of a pathway is evolving that contributes to tissue damage both directly via the formation of peroxynitrite, with its associated toxicities, and indirectly through the amplification of the inflammatory response.
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PMID:Novel roles of nitric oxide in hemorrhagic shock. 1046 45

Aldehyde dehydrogenase from the bioluminescent bacterium, Vibrio harveyi, catalyses the oxidation of long-chain aliphatic aldehydes to acids. The enzyme is unique compared with other forms of aldehyde dehydrogenase in that it exhibits a very high specificity and affinity for the cofactor NADP(+). Structural studies of this enzyme and comparisons with other forms of aldehyde dehydrogenase provide the basis for understanding the molecular features that dictate these unique properties and will enhance our understanding of the mechanism of catalysis for this class of enzyme. The X-ray structure of aldehyde dehydrogenase from V. harveyi has been solved to 2.5-A resolution as a partial complex with the cofactor NADP(+) and to 2. 1-A resolution as a fully bound 'holo' complex. The cofactor preference exhibited by different forms of the enzyme is predominantly determined by the electrostatic environment surrounding the 2'-hydroxy or the 2'-phosphate groups of the adenosine ribose moiety of NAD(+) or NADP(+), respectively. In the NADP(+)-dependent structures the presence of a threonine and a lysine contribute to the cofactor specificity. In the V. harveyi enzyme an arginine residue (Arg-210) contributes to the high cofactor affinity through a pi stacking interaction with the adenine ring system of the cofactor. Further differences between the V. harveyi enzyme and other aldehyde dehydrogenases are seen in the active site, in particular a histidine residue which is structurally conserved with phosphorylating glyceraldehyde-3-phosphate dehydrogenase. This may suggest an alternative mechanism for activation of the reactive cysteine residue for nucleophilic attack.
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PMID:Crystal structure of the NADP+-dependent aldehyde dehydrogenase from Vibrio harveyi: structural implications for cofactor specificity and affinity. 1090 48

The rat luminal endoplasmic-recticulum calcium-binding proteins 1 and 2 (CaBP1 and CaBP2 respectively) are members of the protein disulphide-isomerase (PDI) family. They contain two and three thioredoxin boxes (Cys-Gly-His-Cys) respectively and, like PDI, may be involved in the folding of nascent proteins. We demonstrate here that CaBP1, similar to PDI and CaBP2, can complement the lethal phenotype of the disrupted Saccharomyces cerevisiae PDI gene, provided that the natural C-terminal Lys-Asp-Glu-Leu sequence is replaced by His-Asp-Glu-Leu. Both the in vitro RNase AIII-re-activation assays and in vivo pro-(carboxypeptidase Y) processing assays using CaBP1 and CaBP2 thioredoxin (trx)-box mutants revealed that, whereas the three trx boxes in CaBP2 seem to be functionally equivalent, the first trx box of CaBP1 is significantly more active than the second trx box. Furthermore, only about 65% re-activation of denatured reduced RNase AIII could be obtained with CaBP1 or CaBP2 compared with PDI, and the yield of PDI-catalysed reactions was significantly reduced in the presence of either CaBP1 or CaBP2. In contrast with PDI, neither CaBP1 nor CaBP2 could catalyse the renaturation of denatured glyceraldehyde-3-phosphate dehydrogenase (GAPDH), which is a redox-independent process, and neither protein had any effect on the PDI-catalysed refolding of GAPDH. Furthermore, although PDI can bind peptides via its b' domain, a property it shares with PDIp, the pancreas-specific PDI homologue, and although PDI can bind malfolded proteins such as 'scrambled' ribonuclease, no such interactions could be detected for CaBP2. We conclude that: (1) both CaBP2 and CaBP1 lack peptide-binding activity for GAPDH attributed to the C-terminal region of the a' domain of PDI; (2) CaBP2 lacks the general peptide-binding activity attributed to the b' domain of PDI; (3) interaction of CaBP2 with substrate (RNase AIII) is different from that of PDI and substrate; and (4) both CaBP2 and CaBP1 may promote oxidative folding by different kinetic pathways.
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PMID:Functional roles and efficiencies of the thioredoxin boxes of calcium-binding proteins 1 and 2 in protein folding. 1141 39

The involvement of the lysine residue present at the active site of Ehrlich ascites carcinoma (EAC) cell glyceraldehyde-3-phosphate dehydrogenase (Gra3PDH) was investigated by using the lysine specific reagents trinitrobenzenesulfonic acid (TNBS) and pyridoxal phosphate (PP). Both TNBS and PP inactivated EAC cell Gra3PDH with pseudo-first-order kinetics with the rate dependent on modifier concentration. Kinetic analysis, including a Tsou plot, indicated that both TNBS and PP apparently react with one lysine residue per enzyme molecule. Two of the substrates, d-glyceraldehyde-3-phosphate and NAD, and also NADH, the product and competitive inhibitor, almost completely protected the enzyme from inactivation by TNBS. A comparative study of Gra3PDH of EAC cell and rabbit muscle indicates that the nature of active site of the enzyme is significantly different in these two cells. A double inhibition study using 5,5'-dithiobis(2-nitrobenzoic acid) and TNBS and subsequent reactivation of only the rabbit muscle enzyme by dithiothreitol suggested that a cysteine residue of this enzyme possibly reacts with TNBS. These studies on the other hand, confirm that an essential lysine residue is involved in the catalytic activity of the EAC cell enzyme. This difference in the nature of the active site of EAC cell Gra3PDH that may be related to the high glycolysis of malignant cells has been discussed.
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PMID:Identification of a critical lysine residue at the active site in glyceraldehyde-3-phosphate dehydrogenase of Ehrlich ascites carcinoma cell. Comparison with the rabbit muscle enzyme. 1173 97

Acrolein, a representative carcinogenic aldehyde that could be ubiquitously generated in biological systems under oxidative stress, shows facile reactivity with the epsilon-amino group of lysine to form N(epsilon)-(3-formyl-3,4-dehydropiperidino)lysine (FDP-lysine) as the major product (Uchida, K., Kanematsu, M., Morimitsu, Y., Osawa, T., Noguchi, N., and Niki, E. (1998) J. Biol. Chem. 273, 16058-16066). In the present study, we determined the electrophilic potential of FDP-lysine and established a novel mechanism of protein thiolation in which the FDP-lysine generated in the acrolein-modified protein reacts with sulfhydryl groups to form thioether adducts. When a sulfhydryl enzyme, glyceraldehyde-3-phosphate dehydrogenase, was incubated with acrolein-modified bovine serum albumin in sodium phosphate buffer (pH 7.2) at 37 degrees C, a significant loss of sulfhydryl groups, which was accompanied by the loss of enzyme activity and the formation of high molecular mass protein species (>200 kDa), was observed. The FDP-lysine adduct generated in the acrolein-modified protein was suggested to represent a thiol-reactive electrophile based on the following observations. (i) N(alpha)-acetyl-FDP-lysine, prepared from the reaction of N(alpha)-acetyl lysine with acrolein, was covalently bound to glyceraldehyde-3-phosphate dehydrogenase. (ii) The FDP-lysine derivative reacted with glutathione to form a GSH conjugate. (iii) The acrolein-modified bovine serum albumin significantly reacted with GSH to form a glutathiolated protein. Furthermore, the observation that the glutathiolated acrolein-modified protein showed decreased immunoreactivity with an anti-FDP-lysine monoclonal antibody suggested that the FDP-lysine residues in the acrolein-modified protein served as the binding site of GSH. These data suggest that thiolation of the protein-bound acrolein may be involved in redox alteration under oxidative stress, whereby oxidative stress generates the increased production of acrolein and its protein adducts that further potentiate oxidative stress via the depletion of GSH in the cells.
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PMID:Thiolation of protein-bound carcinogenic aldehyde. An electrophilic acrolein-lysine adduct that covalently binds to thiols. 1203 48


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