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:2.4.2.30 (PARP)
13,611 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

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 ras oncogene products (ras p21s) are 21-KDa proteins with activities of GTP binding and hydrolysis. A number of proteins homologous to ras p21 have been discovered and collectively named small molecular weight GTP-binding proteins. These proteins undergo post-translational modification with isoprenoid residues attached to cysteine in their carboxyl terminal. With this modification, they attach to cellular membranes. The biochemical activities of these proteins, i.e., GTP hydrolysis and binding, are regulated by various regulatory factors such as GDP-GTP exchange proteins and GTPase-activating proteins, but little is known about the cellular functions and physiological pathways through which they regulate these functions. Botulinum C3 ADP-ribosyltransferase, a 23-KDa exoenzyme secreted from certain strains of types C and D Clostridium botulinum, specifically ADP-ribosylates the rho family of these GTP-binding proteins. This ADP-ribosylation occurs at a specific asparagine residue in their putative effector domain, and presumably interferes with their interaction with a putative effector molecule downstream in signal transduction. C3 exoenzyme, when incubated with or microinjected into cultured cells, ADP-ribosylates a rho gene product in the cells, and causes profound cell rounding with loss of adhesion plaques and collapse of stress fiber. Microinjection of an activated mutant of rho A protein, on the contrary, induced extensive adhesion and actin assembly in cultured cells. These results suggest that the rho family of proteins are involved in morphogenesis and motility of cells via assembly and disassembly of cytoskeletal systems, and botulinum ADP-ribosyltransferase is a useful tool for clarifying the molecular mechanism of these processes.
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PMID:[ras oncogene-related small molecular weight GTP-binding protein, rho gene product and botulinum C3 ADP-ribosyltransferase]. 160 29

The role of ADP ribosylation of proteins in the physiological regulation of sporulation in Streptomyces griseus was studied. We report here that both the activity of NAD+: arginine ADP-ribosyltransferase (ADPRT) and the pattern of ADP-ribosylated proteins showed characteristic changes during the life cycle in S. griseus 2682. Analysis off ADP-ribosylated proteins revealed that in a nonsporulating mutant of the parental wild-type (wt) strain (Bld7 mutant), both the activity of ADPRT and the pattern of ADP-ribosylated proteins were different from those of the parental strain. Addition of 3-aminobenzamide (3AB), the most potent inhibitor of ADPRT, inhibited sporulation of S. griseus 2682 and the A-factor (AF)-induced sporulation of S. griseus Bld7, but in both cases the inhibitory effect of 3AB was strictly age-dependent. Using [alpha-32P]GTP, we have demonstrated the presence of GTP-binding proteins in purified cell membranes of S. griseus 2682 and S. griseus Bld7. The same GTP-binding proteins were observed in Bld7 and the wt. AF stimulated the basal GTPase activity of cell membranes of S. griseus 2682 in a concentration-dependent manner, suggesting that GTP-binding proteins might be involved in the AF-induced sporulation process.
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PMID:The possible role of ADP ribosylation in physiological regulation of sporulation in Streptomyces griseus. 161 34

Degranulation of neutrophils involves the differential regulation of the exocytosis of at least two populations of granules. Low molecular weight GTP-binding proteins (LMW-GBPs) have been implicated in the regulation of vesicular traffic in the secretory pathways of several types of cells. In the present study we identify distinct subsets of LMW-GBPs associated with the membranes of neutrophil-specific and azurophilic granules. Ninety-four percent of total [35S]guanosine 5'-(3-O-thio)triphosphate (GTP gamma S) binding activity was equally distributed between the plasma membrane and cytosol with the remaining 6% localized in the granules. In contrast, the cytosol contained only 10% of the total GTPase activity while the specific granules accounted for 13%. [alpha-32P]GTP binding to proteins transferred to nitrocellulose revealed LMW-GBPs in all fractions except the azurophilic granules. The specific granules contained three out of four bands which were found in the plasma membrane; these ranged from 20 to 23 kDa and all were resistant to alkaline extraction. Photoaffinity labeling with [alpha-32P]8-azido-GTP in the presence of micromolar Al3+ identified proteins of 25 and 26 kDa unique to azurophilic granules; these could not be labeled with [alpha-32P]8-azido-ATP and could be extracted by acidic but not alkaline pH. Botulinum C3-mediated [32P]ADP-ribosylation identified proteins of 16, 20, and 24 kDa both in plasma membranes and those of specific granules. An anti-ras monoclonal antibody, 142-24E5, recognized a 20-kDa protein localized to the plasma and specific granule membranes which could not be extracted by alkaline pH, was not a substrate for botulinum C3 ADP-ribosyltransferase, and was translocated from specific granules to plasma membrane after exposure of neutrophils to phorbol myristate acetate. We conclude that neutrophil-specific and azurophilic granules contain distinct subsets of LMW-GBPs which are uniquely situated to regulate the differential exocytosis of these two compartments.
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PMID:Low molecular weight GTP-binding proteins in human neutrophil granule membranes. 189 32

A GTP-binding protein with an Mr of 24,000 was purified from a cholate extract of bovine brain membranes in addition to the previously reported alpha beta gamma-trimeric GTP-binding proteins (G proteins). Partial amino acid sequence analysis of the purified 24-kDa protein revealed that it was not identical to any of the low Mr GTP-binding proteins already reported, but similar to the rac-gene products serving as the substrate of an ADP-ribosyltransferase (C3) purified from the culture medium of Clostridium botulinum type C. However, the 24-kDa protein was not ADP-ribosylated by the botulinum C3 enzyme. The 24-kDa protein was purified as a nucleotide-free form and characterized by the following unique properties distinct from those of alpha beta gamma-trimeric G proteins. (1) Mg2+ was essentially required for nucleotide binding to the 24-kDa protein; there was a progressive increase in its binding affinity for nucleotides as the concentration of the divalent cation was increased. (2) Nucleotides previously bound to the 24-kDa protein were rapidly dissociated from the protein in Mg(2+)-free medium, in accord with the fact that the protein was indeed purified as a nucleotide-free form with Mg(2+)-free solutions. (3) The 24-kDa protein apparently exhibited much lower GTPase activity than do alpha beta gamma-trimeric G proteins because the product GDP was released from the 24-kDa protein in exchange for the substrate GTP only at a very low rate. Based on these findings, a possible role of the 24-kDa protein in cellular signalling is discussed in comparison with well characterized alpha beta gamma-trimeric G proteins.
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PMID:Purification and characterization of a new GTP-binding protein of Mr 24,000 in bovine brain membranes. 190 60

In bovine aortic smooth muscle, about 50% of total GTP-binding activity was present in the cytosol fraction. A major GTP-binding protein (G protein) with a Mr value of about 21,000 (21K G) in this fraction was purified to near homogeneity and characterized. 21K G bound maximally about 0.8 mol of [35S]guanosine 5'-(3-O-thio)triphosphate/mol of protein with a Kd value of about 20 nM. 21K G showed GTPase activity with a turnover number of about 0.007 min-1. 21K G was ADP-ribosylated by botulinum ADP-ribosyltransferase and about 0.4 mol of ADP-ribose was maximally incorporated into 1 mol of 21K G. 21K G and the bovine brain rhoA gene product (rhoA p21) were eluted at the same retention time on C4 reversed-phase high performance liquid chromatography and migrated at the same positions on two-dimensional gel electrophoresis. These results indicate that the major G protein in bovine aortic smooth muscle cytosol is rhoA p21.
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PMID:Identification of a major GTP-binding protein in bovine aortic smooth muscle cytosol as the rhoA gene product. 211 95

We have previously shown that there are multiple GTP-binding proteins (G proteins) with Mr values of about 20,000 in bovine brain membranes and identified one G protein with a Mr of 20,000 as the rho gene product. We have also shown that this rho gene product is ADP-ribosylated by an ADP-ribosyltransferase contaminated in botulinum toxin type C1. In the present studies, we have purified another G protein with a Mr of about 21,000 to near homogeneity from bovine brain membranes by several column chromatographies and identified it as the rhoA gene product. Further analysis of the amino acid sequence of the G protein, which we have purified and identified as the rho gene product previously, has revealed that this G protein is the rhoB gene product. The rhoA gene product binds maximally about 0.9 mol of [35S]guanosine 5'-(3-O-thio) triphosphate (GTP gamma S)/mol of protein with a K d value of about 20 nM. [35S]GTP gamma S-binding to the rhoA gene product is inhibited by pretreatment with N-ethylmaleimide. The rhoA gene product hydrolyzes GTP to liberate Pi with a turnover number of about 0.01 min-1. Moreover, the rhoA gene product is ADP-ribosylated by an ADP-ribosyltransferase contaminated in botulinum toxin type Cl. About 0.3 mol of ADP-ribose is maximally incorporated into 1 mol of the rhoA gene product. The ADP ribosylation of the rhoA gene product does not affect its GTP gamma S-binding or GTPase activity. These properties of the rhoA gene product are similar those of the rhoB gene product described previously. These results together with the earlier observations indicate that there are at least two rho gene products (rhoA, B) among three members of the rho gene family (rhoA, B, C) in bovine brain membranes and that both of them are ADP-ribosylated by an ADP-ribosyltransferase contaminated in botulinum toxin type C1.
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PMID:Purification and characterization from bovine brain membranes of a GTP-binding protein with a Mr of 21,000, ADP-ribosylated by an ADP-ribosyltransferase contaminated in botulinum toxin type C1--identification as the rhoA gene product. 215 99

We reported previously that the ADP-ribosyltransferase in C1 and D botulinum toxins specifically catalyzes ADP-ribosylation of an Mr 22,000 guanine nucleotide-binding protein and that this substrate named Gb (b = botulinum) has an amino acid sequence homologous to that deduced from the rho gene (Narumiya, S., Sekine, A., and Fujiwara, M. (1988) J. Biol. Chem. 263, 17255-17257). In this study we have determined the amino acid sequence at its ADP-ribosylation site. Purified substrate was [32P]ADP-ribosylated by C1 botulinum toxin and digested with trypsin. The radioactive peptides were isolated by reversed-phase high performance liquid chromatography and digested further either with protease V8, with proteases V8 and thermolysin, or with proline endopeptidase and thermolysin. By this procedure three radioactive peptides were obtained, and their amino acid sequences were X-Tyr-Val-Ala-Asp-Ile-Glu, X-Tyr, and Val-Phe-Glu-X-Tyr in which no amino acid peak was found in X. During the sequencing the radioactivity quantitatively adhered to the sequencing filter and was not eluted with either of the identified amino acid residues. Analysis of the protein without the ADP-ribosylation yielded the corresponding sequence as Thr-Val-Phe-Glu-Asn-Tyr which corresponds to Thr37-Tyr42 in the amino acid sequence deduced from the Aplysia rho gene. These results strongly suggest that the asparagine residue is the ADP-ribosylation site in the rho gene product. This ADP-ribose protein bond was stable in 0.5 M hydroxylamine at pH 7.5 at 37 degrees C for at least 5 h. The ADP-ribosylation of this protein affected neither its GTPase- nor its [35S]guanosine 5'-O-thiotriphosphate-binding activity.
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PMID:Asparagine residue in the rho gene product is the modification site for botulinum ADP-ribosyltransferase. 249 16

The bacterial toxins, choleragen and pertussis toxin, inhibit the light-stimulated GTPase activity of bovine retinal rod outer segments by catalysing the ADP-ribosylation of the alpha-subunit (T alpha) of transducin [Abood, Hurley, Pappone, Bourne & Stryer (1982) J. Biol. Chem. 257, 10540-10543; Van Dop, Yamanaka, Steinberg, Sekura, Manclark, Stryer & Bourne (1984) J. Biol. Chem. 259, 23-26]. Incubation of retinal rod outer segments with NAD+ and a purified NAD+:arginine ADP-ribosyltransferase from turkey erythrocytes resulted in approx. 60% inhibition of GTPase activity. Inhibition was dependent on both enzyme and NAD+, and was potentiated by the non-hydrolysable GTP analogues guanosine 5'-[beta gamma-imido]triphosphate (p[NH]ppG) and guanosine 5'-[beta gamma-methylene]triphosphate (p[CH2]ppG). The transferase ADP-ribosylated both the T alpha and T beta subunits of purified transducin. T alpha (39 kDa), after ADP-ribosylation, migrated as two distinct peptides with molecular masses of 42 kDa and 46 kDa on SDS/polyacrylamide-gel electrophoresis. T beta (36 kDa), after ADP-ribosylation, migrated as a 38 kDa peptide. With purified transducin subunits, it was observed that the GTPase activity of ADP-ribosylated T alpha, reconstituted with unmodified T beta gamma and photolysed rhodopsin, was decreased by 80%; conversely, reconstitution of T alpha with ADP-ribosyl-T beta gamma resulted in only a 19% inhibition of GTPase. Thus ADP-ribosylation of T alpha, the transducin subunit that contains the guanine nucleotide-binding site, has more dramatic effects on GTPase activity than does modification of the critical 'helper subunits' T beta gamma. To elucidate the mechanism of GTPase inhibition by transferase, we studied the effect of ADP-ribosylation on p[NH]pp[3H]G binding to transducin. It was shown previously that modification of transducin by choleragen, which like transferase ADP-ribosylates arginine residues, did not affect guanine nucleotide binding. ADP-ribosylation by the transferase, however, decreased p[NH]pp[3H]G binding, consistent with the hypothesis that choleragen and transferase inhibit GTPase by different mechanisms.
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PMID:Inhibition of the GTPase activity of transducin by an NAD+:arginine ADP-ribosyltransferase from turkey erythrocytes. 282 39

Recently we demonstrated the presence in calf thymocytes of a GTP-binding protein (G-protein) composed of three polypeptides, 54, 41, and 27 kDa, which was physically and functionally associated with a soluble phosphoinositides-specific phospholipase C (PI-phospholipase C). The properties of this G protein were further investigated with the following results. 1) In addition to the ability to bind [35S]guanosine-5'-[gamma-thio]triphosphate (GTP gamma S), the G-protein exhibited GTPase activity, which was enhanced by Mg2+, phosphatidylethanolamine, phosphatidylserine, and phosphatidylinositol, but inhibited by sodium cholate, GTP gamma S and F-.2) The 54-kDa polypeptide was ADP-ribosylated by pertussis toxin and also by endogenous membrane-bound ADP-ribosyltransferase, but none of these three polypeptides was ADP-ribosylated by cholera toxin. 3) The G-protein did not cross-react with either anti-rat brain alpha 1 (alpha-subunit of inhibitory G-protein, G1), alpha 0 (alpha-subunit of other G1-like G-protein, G0) or beta gamma antibodies. 4) Incubation of this G Protein with GTP gamma S caused dissociation of the three polypeptides. 5) The 27 kDa polypeptide showed GTP-binding activity and enhanced the phosphatidylinositol 4,5-bisphosphate hydrolysis by purified PI-phospholipase C. These results suggest that the PI-phospholipase C-associated G-protein in calf thymocytes may be a novel one and that it is involved in the regulation of PI-phospholipase C activity.
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PMID:Properties of a novel GTP-binding protein which is associated with soluble phosphoinositides-specific phospholipase C. 283 52


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