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Query: UMLS:C0043167 (pertussis)
 19,595 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Bordetella pertussis, the causative agent of whooping cough, releases pertussis toxin in an inactive form. The toxin consists of an A protomer containing one S1 peptide subunit and a B oligomer containing several other peptide subunits. The toxin binds to cells via the B oligomer, and the S1 subunit is activated and expresses ADP-ribosyltransferase and NAD glycohydrolase activities. Treatment of purified toxin with dithiothreitol (DTT) in vitro increases both activities. ATP and the detergent 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) synergistically reduce the A0.5 (activation constant) for DTT from greater than 100 mM to 200 microM. We studied the structure-activity relationships of activators of the toxin. In the presence of CHAPS (1%) and DTT (10 mM) the following compounds increased the NAD glycohydrolase activity of the toxin with the following A0.5's in microM and fraction of the ATP effect in parentheses: ATP, 0.2 (1.0); ADP, 6 (0.8); UTP, 15 (0.7); GTP, 35 (0.6); pyrophosphate, 45 (0.7); triphosphate, 60 (0.6); tetraphosphate, greater than or equal to 170 (greater than or equal to 0.4). Thus, the polyphosphate moiety is sufficient to stimulate the toxin, and the adenosine moiety confers upon ATP its extraordinary affinity for the toxin. Phospholipid and detergents could substitute for CHAPS in the activation of the toxin. Glutathione substituted for DTT with an A0.5 of 2 mM, a concentration within the range found in eucaryotic cells. Thus, membrane lipids and cellular concentrations of glutathione and ATP are sufficient to activate pertussis toxin without the need for a eucaryotic enzymatic process.
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PMID:Structure-activity analysis of the activation of pertussis toxin. 303 Mar 99

15-Hydroxyeicosatetraenoic acid [15-(S)-HETE], a major arachidonic acid metabolite produced from the 15-lipoxygenase pathway, has been characterized as an antiinflammatory cellular mediator since it can inhibit the in vivo and in vitro formation of the proinflammatory leukotrienes via the 5-lipoxygenase pathway in various cells. 15-HETE has been confirmed to inhibit the 5-lipoxygenase in rat basophilic leukemia cell (RBL-1) homogenates with an I50 = 7.7 microM. The I50 of the 12-HETE isomer was 6 microM whereas prostaglandin F2 alpha was ineffective. In order to examine the mechanistic basis underlying the inhibitory action of 15-HETE, association assays of [3H]-15-HETE with RBL-1 subcellular fractions were carried out. The presence of the zwitterionic detergent CHAPS enhanced specific [3H]-15-HETE binding in the membrane fractions three-fold and specific 15-HETE binding was distributed among the nuclear (32%)-, granule (19%)-, plasma membrane (35%)-, and cytosol (14%)-enriched fractions. Studies using combined granule and plasma membrane enriched-, CHAPS treated-fractions showed that [3H]-15-HETE binding was time-dependent, specific and reversible, sensitive to pertussis toxin treatment, and indicated a single class of binding sites with a Kd = 460 +/- 160 nM and Bmax = 5.0 +/- 1.1 nM. Competition experiments showed that the order of 15-HETE or analogs in inhibiting the binding of [3H]-15-HETE was: 15(S)-HETE > or = 12-(S)-HETE = 5-(S)-HETE > 15-(R)-HETE > arachidonic acid. Prostaglandin F2 alpha and lipoxin B4 were ineffective as competitors. The similar profiles of the binding assays and inhibition of the 5-lipoxygenase suggest that 15-HETE binding sites may mediate this inhibitory action of 15-HETE.
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PMID:Characterization of specific subcellular 15-hydroxyeicosatetraenoic acid (15-HETE) binding sites on rat basophilic leukemia cells. 778 91

Pertussis toxin and adenylate cyclase toxin both contribute to the pathogenesis of whooping cough. Production of these proteins is controlled by the bvg locus, which is inactive at 25 degrees C, but at 37 degrees C produces a Vir+ phenotype. In view of the temperature dependence of virulence factor synthesis, the effects of temperature and host factors on their action were examined. The NAD glycohydrolase activity of the S1 subunit of pertussis toxin was enhanced by CHAPS, a zwitterionic detergent, with a temperature optimum of approximately 35 degrees C. Similar temperature optima for the ADP-ribosylation by pertussis toxin of transducin and recombinant Go alpha were observed. Since the temperature--activity relationship of S1 differed from that of S1 in activated holotoxin, and since S1 in activated holotoxin was more stable at 42 degrees C than was S1, it appears that S1 associated with the B oligomer components may, in fact, be an active species. Bordetella pertussis adenylate cyclase is activated by a host factor, calmodulin. In the absence of calmodulin, the temperature optimum for enzymatic activity was approximately 25 degrees C, whereas in its presence it was approximately 35 degrees C. Thus, the temperature optima for pertussis and adenylate cyclase toxins, virulence factors whose production is increased through the bvg locus at physiological temperatures, are either at or near these temperatures when stimulated by host factors.
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PMID:Effect of temperature and host factors on the activities of pertussis toxin and Bordetella adenylate cyclase. 780 92

CHAPS-solubilized muscarinic receptors (mAChRs) retain selective agonist or antagonist binding and sensitivity to GTP and ADP-ribosylation by pertussis toxin that ribosylates a group of G protein called Gi. This suggests that CHAPS solubilizes the mAChR-Gi complex both from neonatal and adult neurons. The gradient centrifugation of solubilized samples indicated that the neonatal samples contain mostly Gi-coupled mAChR (mAChRn) exhibiting a high affinity for [3H]OxM, but the adult samples contain a comparable quantity of Gi coupled mAChR (mAChRa1) and uncoupled mAChRs (mAChRa2) exhibiting low affinity for the agonist. The binding of [3H]OxM to or the dissociation of [3H]OxM from mAChRn and mAChRa1 was sensitive to GTP. However, the effects of GTP were sensitive to Na+ for mAChRn but not for mAChRa1. ADP-ribosylation by pertussis toxin but not by cholera toxin abolished the effects of Na+ on agonist binding to solubilized mAChRn, suggesting that NaCl affects mAChRn not by interacting with the receptor but by interacting with the mAChR-Gi complex. The subtype composition of mAChRn, mAChRa1 and mAChRa2 was studied by determining the IC50 and the Ki values for the inhibition by subtype-selective antagonists in the binding of [3H]OxM to the receptors. The neonatal and adult samples exhibited differences in the distribution of the mAChR subtypes, the neonatal samples containing mostly the M1-mAChR subspecies but the adult samples containing M1-, M2- and M3-mAChR subspecies. The pirenzepine-induced inhibition in the binding of [3H]OxM both to M1-mAChRn and M1-mAChRa1 was sensitive to GTP. The GTP-induced effects on mAChRn were modulated by Na+, but the effect on M1-mAChRa1 was not. This suggests functional differences between Gi coupled M1-mAChRn and M1-mAChRa1.
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PMID:Regulation of the CHAPS-solubilized muscarinic receptors by an inhibitory GTP binding protein (Gi) in the brain of neonatal and adult rats. 795 18

The present investigation was undertaken to characterize cannabinoid receptor binding in the absence of the membrane environment, inasmuch as cannabinoid drugs have been noted to influence the behavior of integral membrane proteins. The zwitterionic detergent 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) was able to solubilize the cannabinoid receptor from rat brain membranes, with the greatest yield and specific activity being obtained at a detergent/protein ratio of 0.5:1. [3H]CP-55940 bound to a single class of binding sites in the CHAPS extract, which exhibited a Kd of 0.94 nM as determined by nonlinear regression analysis of equilibrium binding data. The order of potency for cannabinoid agonists in heterologous equilibrium binding studies was CP-55244 > or = desacetyllevonantradol > delta 9-tetrahydrocannabinol > cannabinol >> cannabidiol, consistent with the relative affinities for these agonists in brain membrane preparations. CP-55243, the biologically inactive enantiomer of CP-55244, competed for binding of [3H]CP-55940 by < 50% at 1 microM, similar to its poor affinity for the receptor in membranes. The CHAPS-solubilized cannabinoid receptor exhibited functional interactions with guanine nucleotide-binding proteins (G proteins). GTP and nonhydrolyzable analogs decreased [3H]CP-55940 binding by 75%. The concentration-effect curves for guanine nucleotides exhibited a potency order similar to that observed for other G protein-linked receptors. Kinetic analyses indicated that GTP analogs increased the rate of agonist dissociation, decreasing the t1/2 from 60 min at 0-4 degrees to a multiphasic dissociation that exhibited a component having a t1/2 of < 1 min. The cannabinoid agonist desacetyllevonantradol was able to reduce pertussis toxin-catalyzed ADP-ribosylation of G proteins by 50%, demonstrating a receptor effect on G protein functions. These studies demonstrate that the membrane environment is not necessary for agonist binding to the cannabinoid receptor. Furthermore, the cannabinoid receptor maintains its functional interactions with pertussis toxin-sensitive G proteins in detergent solution.
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PMID:Solubilization of the cannabinoid receptor from rat brain and its functional interaction with guanine nucleotide-binding proteins. 842 66


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