Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.4.2.30 (PARP)
 13,611 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Highly purified, polymyxin-released, low molecular weight Escherichia coli heat-labile enterotoxin (LT) catalyzed the hydrolysis of NAD to ADP-ribose and nicotinamide. This NAD glycohydrolase activity was stimulated by dithiothreitol and was independent of cellular components. Nicotinamide formation was enhanced by arginine methyl ester > d-arginine congruent with l-arginine congruent with guanidine. A 20-fold increase in activity was noted with arginine methyl ester, and maximal activity again required dithiothreitol. When the reaction was initiated with toxin, a delay was observed before a constant rate was established. The reaction products found after incubation of [adenine-U-(14)C]NAD and l-[(3)H]arginine or unlabeled arginine methyl ester with the enterotoxin had mobilities on thin-layer chromatograms similar to the reaction products obtained after incubation of choleragen with these substrates and are consistent with the formation of ADP-ribose-l-arginine and ADP-ribose-l-arginine methyl ester, respectively. Both toxins, which catalyze the NAD-dependent activation of adenylate cyclase, thus appear to possess NAD glycohydrolase and ADP-ribosyltransferase activities. Although the activities of both toxins are dependent on dithiothreitol, Escherichia coli enterotoxin exhibited optimal activity in Tris (Cl(-)) (pH 7.5) and was inhibited by high concentrations of potassium phosphate (pH 7.0) or low pH (sodium acetate, pH 6.2). It appears that the optimal assay conditions as well as the kinetic constants for the reactants differ from those previously noted with choleragen. It is probable therefore that although the two toxins catalyze similar reactions, they differ in primary structure. The presence of transferase and glycohydrolase activities in structurally distinct toxins that activate adenylate cyclase strengthens our hypothesis that the ADP-ribosylation of arginine is a model for the NAD-dependent activation of adenylate cyclase; activation may result from ADP-ribosylation of the cyclase itself or of a protein that regulates its activity.
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PMID:Activation of adenylate cyclase by heat-labile Escherichia coli enterotoxin. Evidence for ADP-ribosyltransferase activity similar to that of choleragen. 20 60

Pseudomonas aeruginosa exoenzyme S is an adenosine diphosphate ribosyltransferase distinct from Pseudomonas toxin A. Exoenzyme S catalyzes the transfer of radioactivity from all portions of radiolabeled NAD+ except nicotinamide. Digestion of the radiolabeled product(s) formed in the presence of [adenine-14C]NAD+ and exoenzyme S with snake venom phosphodiesterase yields only AMP, suggesting that ADP-ribose is present as monomers and not as poly(ADP-ribose). Exoenzyme S does not catalyze the transfer of ADP-ribose from NAD+ to elongation factor 2, as do toxin A and diphtheria toxin, but to one or more other proteins present in crude extracts of wheat germ or rabbit reticulocytes and in partially purified preparations of elongation factor I. The ADP-ribosyltransferase activity of exoenzyme S is distinct from toxin A by several tests: it is not neutralized by toxin A antibody, it is destroyed rather than potentiated by pretreatment with urea, and it is more heat stable. These latter observations and the substrate specificity suggest that exoenzyme S is different from any previously described prokaryotic ADP-ribosyltransferase.
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PMID:Pseudomonas aeruginosa exoenzyme S: an adenosine diphosphate ribosyltransferase distinct from toxin A. 21 Apr 53

An ADP-ribosyltransferase was purified approximately 500-fold from the supernatant fraction of turkey erythrocytes. The enzyme hydrolyzed [carbonyl-(14)C]NAD to ADP-ribose and [carbonyl-(14)C]nicotinamide at a low rate. Nicotinamide formation from NAD was enhanced by arginine methyl ester > D-arginine approximately L-arginine > guanidine; lysine, histidine, and citrulline were ineffective. Incubation of [adenine-U-(14)C]NAD and arginine methyl ester or arginine with the purified enzyme resulted in the formation of new compounds that contained (14)C, reacted with ninhydrin, and quenched background fluorescence of thin-layer plates viewed in ultraviolet light. Their mobilities on thin-layer chromatograms were indistinguishable from those of ADP-ribosylarginine methyl ester and ADP-ribosylarginine formed during incubation of choleragen with NAD and arginine methyl ester or arginine, respectively [Moss, J. & Vaughan, M. (1977) J. Biol. Chem. 252, 2455-2457]. The purified transferase also catalyzed the incorporation of label from [adenine-(14)C]-NAD into lysozyme, histones and polyarginine. When the (14)C-labeled lysozyme was incubated with snake venom phosphodiesterase, the radioactivity was released and, on thin-layer chromatograms, exhibited a mobility indistinguishable from that of 5'-AMP, as would be expected of an ADP-ribosylated protein, but not of a poly(ADP-ribosylated) product. The purified transferase activated rat brain adenylate cyclase and, as is the case with choleragen, activation was absolutely dependent on NAD. The presence in the avian erythrocyte of a protein that, like choleragen and Escherichia coli heat-labile enterotoxin, apparently activates adenylate cyclase and possesses ADP-ribosyl transferase activity is consistent with the view that the mechanisms through which the bacterial toxins produce pathology are not entirely foreign to vertebrate cells, at least some of which may possess and employ an analogous mechanism for activation of adenylate cyclase.
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PMID:Isolation of an avian erythrocyte protein possessing ADP-ribosyltransferase activity and capable of activating adenylate cyclase. 21 2

Choleragen exerts its effect on cells through activation of adenylate cyclase. Choleragen initially interacts with cells through binding of the B subunit of the toxin to the ganglioside GM1 on the cell surface. Subsequent events are less clear. Patching or capping of toxin on the cell surface may be an obligatory step in choleragen action. Studies in cell-free systems have demonstrated that activation of adenylate cyclase by choleragen requires NAD. In addition to NAD, requirements have been observed for ATP, GTP, and calcium-dependent regulatory protein. GTP also is required for the expression of choleragen-activated adenylate cyclase. In preparations from turkey erythrocytes, choleragen appears to inhibit an isoproterenol-stimulated GTPase. It has been postulated that by decreasing the activity of a specific GTPase, choleragen would stabilize a GTP-adenylate cyclase complex and maintain the cyclase in an activated state. Although the holotoxin is most effective in intact cells, with the A subunit having 1/20th of its activity and the B subunit (choleragenoid) being inactive, in cell-free systems the A subunit, specifically the A1 fragment, is required for adenylate cyclase activation. The B protomer is inactive. Choleragen, the A subunit, or A1 fragment under suitable conditions hydrolyzes NAD to ADP-ribose and nicotinamide (NAD glycohydrolase activity) and catalyzes the transfer of the ADP-ribose moiety of NAD to the guandino group of arginine (ADP-ribosyltransferase activity). The NAD glycohydrolase activity is similar to that exhibited by other NAD-dependent bacterial toxins (diphtheria toxin, Pseudomonas exotoxin A), which act by catalyzing the ADP-ribosylation of a specific acceptor protein. If the ADP-ribosylation of arginine is a model for the reaction catalyzed by choleragen in vivo, then arginine is presumably an analog of the amino acid which is ADP-ribosylated in the acceptor protein. It is postulated that choleragen exerts its effects on cells through the NAD-dependent ADP-ribosylation of an arginine or similar amino acid in either the cyclase itself or a regulatory protein of the cyclase system.
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PMID:Mechanism of action of choleragen. 21 41

The fibroblast cell line L929 contains a constitutively expressed NO synthase (EC 1.14.29.-) activity, which can be increased about 10-fold by tumour-necrosis factor alpha (TNF-alpha). Activities of the constitutive and the inducible enzymes are tetrahydrobiopterin-independent and can be inhibited by L-NG-nitroarginine. Induction of NO synthase by TNF-alpha was prevented by inhibitors of poly(ADP-ribose) polymerase, namely nicotinamide, 3-methoxybenzamide and 3-aminobenzamide. TNF-alpha did not lead to an increase in ADP-ribosyltransferase activity nor to a change in the pattern of ADP-ribosylated proteins. The inhibitors were only active during the first 4-5 h after exposure to TNF-alpha and they were found to suppress synthesis of protein, DNA and RNA. These data suggest that the inhibitors prevent induction of NO synthase by interference with RNA and protein synthesis. It is not yet known which reactions of these biosynthetic processes are affected by the inhibitors.
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PMID:Induction of nitric oxide synthase in L929 cells by tumour-necrosis factor alpha is prevented by inhibitors of poly(ADP-ribose) polymerase. 128 Jan 12

NG108-15 neuroblastoma x glioma somatic hybrid cells were permeabilized in the presence of [32P]NAD+ and then cultured for 18 h. Resolution of the cell proteins on polyacrylamide gels revealed [32P]ADP-ribosylation of five major protein species with molecular mass values of 52 kDa, 44 kDa, 35 kDa, 30 kDa and 25 kDa. A similar pattern of labelling was also seen when NG108-15 cell membranes were incubated with [32P]NAD+ and hydrolysis of the product revealed mono(ADP-ribosyl)ation. Immunoprecipitation of these products with anti-Gs alpha antiserum revealed a single band identical to cholera toxin substrate. Culture of [32P]NAD(+)-loaded cells for 18 h in the presence of 50 mM-nicotinamide inhibited the eukaryotic mono(ADP-ribosyl)transferase activity. Inhibition of the eukaryotic enzyme was also accompanied by an increase in the abundance of Gs alpha, whether measured by Western blotting with anti-Gs alpha antibody (two separate antisera) or by cholera toxin-dependent [32P]ADP-ribosylation. There was no accompanying change in the abundance of G beta. The increase in Gs alpha abundance in nicotinamide-treated NG108-15 cells was accompanied by a 2-fold increase in basal adenylate cyclase activity (measured in the presence of GTP), and by a smaller but significant increase in iloprost-dependent activation of adenylate cyclase. Receptor number or affinity was not affected by nicotinamide, since this treatment did not alter the binding parameters of [3H]iloprost to NG108-15 cell membranes. Short-term exposure of cells to nicotinamide for 1 h revealed no significant difference in either basal or agonist-stimulated adenylate cyclase activity. These results reveal that mono(ADP-ribosyl)ation of Gs alpha by eukaryotic ADP-ribosyltransferase modifies the abundance and activity of Gs alpha in NG108-15 cells, and hence may play a role in the hormonal regulation of cell function.
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PMID:Gs alpha is a substrate for mono(ADP-ribosyl)transferase of NG108-15 cells. ADP-ribosylation regulates Gs alpha activity and abundance. 128 Jan 14

A critical component of immune responsiveness is the localization of effector cells at sites of inflammatory lesions. Adhesive molecules that may play a role in this process have been described on the surfaces of both lymphocytes and connective tissue cells. Adhesive interactions of T lymphocytes with fibroblasts or endothelial cells can be inhibited by preincubation of the fibroblasts or endothelial cells with antibody to intercellular adhesion molecule 1 (CD54) or by preincubation of the T cells with antibody to lymphocyte function-associated Ag 1 (CD11a/CD18), molecules shown to be important in several other cell-cell adhesive interactions. Here we show that gamma-irradiation of human T lymphocytes impaired their ability to adhere to both fibroblasts and endothelial cells. This impairment was not associated with a loss of cell viability or of cell surface lymphocyte function-associated Ag 1 expression. gamma-Irradiation of T cells is known to result in the activation of ADP-ribosyltransferase, an enzyme involved in DNA strand-break repair, causing subsequent depletion of cellular nicotinamide adenine dinucleotide (NAD) pools by increasing NAD consumption for poly(ADP-ribose) formation. Preincubation of T cells with either nicotinamide or benzamide [corrected], both known inhibitors of ADP-ribosyltransferase, completely reversed the suppressive effects of gamma-irradiation on T cell adhesion. The maintenance of adhesion was accompanied by inhibition of irradiation-induced depletion of cellular NAD. These experiments suggest that the impairment of cellular immune function after irradiation in vivo may be caused, in part, by defective T cell emigration and localization at inflammatory sites.
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PMID:Impairment of lymphocyte adhesion to cultured fibroblasts and endothelial cells by gamma-irradiation. 134 88

Transducin is the retinal rod outer segment (ROS)-specific G protein coupling the photoexcited rhodopsin to cyclic GMP-phosphodiesterase. The alpha subunit of transducin is known to be ADP-ribosylated by bacterial toxins. We investigated the possibility that transducin is modified in vitro by an endogenous ADP-ribosyltransferase activity. By using either ROS, cytosolic extract of ROS or purified transducin in the presence of [alpha-32P]nicotinamide adenine dinucleotide (NAD+), the alpha and beta subunits of transducin were found to be radiolabeled. The labeling was decreased by snake venom phosphodiesterase I (PDE I). The modification was shown to be mono ADP-ribosylation by analyses on thin layer chromatography of the PDE I-hydrolyzed products which revealed only 5'AMP residues. In addition we report that sodium nitroprusside activates the ADP-ribosylation of transducin.
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PMID:Mono ADP-ribosylation of transducin catalyzed by rod outer segment extract. 151 16

C3 ADP-ribosyltransferase is an exoenzyme produced by certain strains of Clostridium botulinum types C and D, which specifically ADP-ribosylates rho proteins in eukaryotic cells. Using the photoaffinity probe [alpha-32P]nicotinamide-2-azidoadenine dinucleotide, we have identified the adenine ring binding domain of the NAD+ binding site. The specificity of labeling was demonstrated by saturation effects and protection by the natural compound at physiologically relevant concentrations. Saturation of labeling was observed at 50 microM. Protection experiments indicated an 80% protection of labeling by 100 microM NAD+ when protein was photolyzed in the presence of 10 microM probe. Trypsin or Staphylococcus aureus V8 protease digestion of the photolabeled protein, along with boronate affinity chromatography and immobilized metal affinity chromatography, was used to specifically isolate the peptide region photolabeled with the probe. The peptide corresponded to Phe9-Gly19 near the N terminus.
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PMID:NAD+ binding site of Clostridium botulinum C3 ADP-ribosyltransferase. Identification of peptide in the adenine ring binding domain using 2-azido NAD. 163 27

Stimulating bone marrow derived macrophages with LPS results in the induction of NO-synthase as measured by NO2- formation. Inhibitors of poly(ADP-ribose)polymerase, namely nicotinamide, 3-aminobenzamide and 3-methoxybenzamide, prevented NO2- formation in a dose dependent manner. Inhibition was most effective if the inhibitors were added at the same time as LPS. When added 10 h after exposure to LPS, a time at which expression of the enzyme had reached its maximum, no inhibition was observed. The inhibitors also blocked early events in activation such as protein and RNA-synthesis as well as DNA-synthesis. Thus prevention of NO2- formation may be related to inhibition of these events. Activation of macrophages by LPS was not accompanied by an increase but rather by a small decrease in ADP-ribosyltransferase activity. Whether this decrease plays a physiological role in activation needs further exploration.
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PMID:Inhibitors of poly (ADP-ribose) polymerase suppress lipopolysaccharide-induced nitrite formation in macrophages. 171 89


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