UMLS:C0043167 - FACTA Search

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

Computer analysis of the three-dimensional structure of ADP-ribosylating toxins showed that in all toxins the NAD-binding site is located in a cavity. This cavity consists of 18 contiguous amino acids that form an alpha-helix bent over a beta-strand. The tertiary folding of this structure is strictly conserved despite the differences in the amino acid sequence. Catalysis is supported by two spatially conserved amino acids, each flanking the NAD-binding site. These are: a glutamic acid that is conserved in all toxins, and a nucleophilic residue, which is a histidine in the diphtheria toxin and Pseudomonas exotoxin A, and an arginine in the cholera toxin, the Escherichia coli heat-labile enterotoxins, the pertussis toxin and the mosquitocidal toxin of Bacillus sphaericus. The latter group of toxins presents an additional histidine that appears important for catalysis. This structure suggests a general mechanism of ADP-ribosylation evolved to work on different target proteins.
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PMID:Common features of the NAD-binding and catalytic site of ADP-ribosylating toxins. 783 May 59

The unmasking of the low concentration effect of angiotensin II (AII) was identified within the concentration ranges of 10(-13) to 10(-11) M of AII by PD 121981 (5-diphenylacetyl-1-(4-methoxy-3-methylbenzyl)- 4,5,6,7-tetrahydro-1H-imidazo[4,5-c]-pyridine-6-carboxylic acid) and 10(-12) to 3 x 10(-10) M of AII by CGP 42112 (nicotinic acid-Tyr-(N alpha-benzyl-oxycarbonyl-Arg)Lys-His-Pro-IIe-OH), AT2 antagonists, in association with the ordinary contraction curve, i.e., high concentration effect (at 3 x 10(-10)-10(-6) M of AII), in the rabbit abdominal aorta. Thus, they showed clear biphasic features of AII-induced contraction curves. However, this was not the case for angiotensin I and angiotensin III. This PD 121981-evoked low concentration effect of AII was selectively inhibited by DuP 753 (0.01-1 nM), dithiothreitol (10 and 100 microM), pertussis toxin (50 and 300 ng/ml, for 2 hr), nifedipine (1 and 10 microM) and 8-(diethylamino)octyl 3,4,5-trimethoxybenzoate hydrochloride (1 and 3 microM), which suggests the receptors were the AT1 subtype. However, the high concentration effect of AII was not affected by these drugs within the concentration ranges used in the present studies. These myographic results were almost consistent with the features of the intracellular Ca++ changes. Thus, it was concluded that the receptors that mediate the low concentration effect of AII belong to the AT1 subtype. However, the current study did not determine the mechanism by which PD 121981 and CGP 42112 evoked the up-regulation of the AT1 receptors.
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PMID:Characterization of PD 121981- and CGP 42112-induced unmasking of low concentration effects of angiotensin II in rabbit abdominal aorta. 799 73

Pertussis toxin is a member of ADP-ribosylating bacterial toxins that are capable of catalyzing the cleavage of the N-glycosidic bond of NAD+ and the transfer of its ADP-ribose moiety to G proteins. The catalytic S1 subunit of pertussis toxin uses signal transducing G proteins as acceptor substrates but can also catalyze the transfer of the ADP-ribose moiety to water in the absence of G proteins. Site-directed mutagenesis followed by kinetic analyses of truncated soluble mutant proteins revealed that His-35 of S1 is a catalytic residue because alterations of this residue affect the turnover rate of NAD-glycohydrolysis by approximately two orders of magnitude without significantly affecting substrate binding. Replacement of the imidazole of His-35 by the side chain of glutamine maintained the highest residual activity. The pH dependence of the enzyme activity showed only slight variations over the experimental range with an optimum at pH 7.5 and an approximate pKa of 6.5 to 7. This pH dependence was abolished by the Gln substitution, which still retained significant activity, suggesting that His-35 probably does not act as a true base but rather as a proton acceptor. Direct catalytic roles for several other residues were ruled out. Ser-52 substitutions resulted in slight alterations of both kcat and Km for NAD+ suggesting an involvement in maintaining the local geometry of the active site rather than a direct role in catalysis for this residue. Kinetic studies on mutants with substitutions of Ser-40 indicate a role in NAD+ binding for this residue. In conjunction with previous findings, these studies suggest that the NAD-glycohydrolase activity of S1 utilizes 2 catalytic residues, His-35 and the previously identified Glu-129. The enzyme mechanism could therefore proceed through an activation by polarization of the acceptor substrate water or G protein by His-35, and the stabilization of an oxocarbonium-like transition state intermediate by Glu-129.
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PMID:The NAD-glycohydrolase activity of the pertussis toxin S1 subunit. Involvement of the catalytic HIS-35 residue. 811 96

Molecular modeling of the S1 subunit (S1) of pertussis toxin with other ADP-ribosylating bacterial exotoxins predicted that histidine 35 (His-35) would residue within the active site of S1. Recombinant derivatives of S1 (rS1 and the C180 peptide) which contained either a H35Q or H35P mutation were analyzed to determine the role of His-35 in ADP-ribosylation. C180 peptide is a recombinant peptide composed of the amino-terminal 180 amino acids of S1. Under linear velocity conditions, C180H35Q possessed 2% of wild type C180 peptide activity and C180H35P possessed no detectable activity in the ADP-ribosylation of transducin. The H35Q mutation did not change the affinity of recombinant peptides for NAD or two targets for ADP-ribosylation, transducin, or alpha i3C20, but did lower the kcat in the NAD glycohydrolase and ADP-ribosyltransferase reactions. Neither the H35Q nor H35P mutation reduced the ability of recombinant proteins to be photocross-linked with NAD which was consistent with the His-35 mutations not reducing the affinity for NAD. These data indicate that His-35 does not reduce the affinity of S1 for NAD or transducin, but functions as a catalytic residue in the ADP-ribosylation reaction possibly in a hydrogen bonding capacity.
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PMID:Role of histidine 35 of the S1 subunit of pertussis toxin in the ADP-ribosylation of transducin. 814 93

Molecular modeling and alignment of the primary amino acid sequence of the S1 subunit (S1) of pertussis toxin (PT) with other members of the family of ADP-ribosylating bacterial exotoxins predicted that tyrosine-98 (Y98) of S1 was a conserved residue among these exotoxins. To extend our understanding of the relationship between pertussis toxin and the other ADP-ribosylating exotoxins, we defined the function of Y98 of S1. Using site-directed mutagenesis, Y98 of S1 was substituted with alanine (Y98A), leucine (Y98L), histidine (Y98H), and phenylalanine (Y98F). Mutations were analyzed in the C180 peptide and C219 peptide, recombinant derivatives of S1 which contain the first 180 and 219 amino-terminal residues of S1, respectively. Periplasmic extracts containing the Y98n peptides expressed similar specific activities for the ADP-ribosylation of transducin (Gt) as the periplasmic extract containing wild-type peptides. Mutations at Y98 influenced the subcellular localization of the respective Y98n peptide. The majority of the wild-type Y98 and Y98F peptides localized to the periplasmic extract, while the majority of Y98A and Y98L peptides were associated with the insoluble bacterial outer membrane. Purified C180Y98A and C180Y98F and partially purified C180Y98H peptides possessed similar specific activities for the ADP-ribosylation of Gt as the wild-type C180 peptide. KmNAD and kcat for C180Y98A and C180Y98F in the NAD glycohydrolase reaction were similar to the wild-type C180 peptide. These data show that the R group of Y98 does not participate in the ADP-ribosylation of Gt, but appears to contribute to the proper folding of S1.
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PMID:Biochemical analysis of mutations at tyrosine-98 of the S1 subunit of pertussis toxin. 831 78

Pertussis toxin is a complex protein composed of five different subunits, named S1 through S5 and arranged in an A-B structure. The B oligomer, composed of S2 through S5, is the receptor-binding moiety, and the A promoter, composed of S1, is the enzymatically active moiety. S1 catalyzes the ADP-ribosylation of a cysteine in the alpha subunit of heterotrimeric G proteins. In the absence of G proteins it also catalyzes the cleavage of NAD+ into ADP-ribose and nicotinamide. Molecular dissection has indicated that the C-terminal domain of S1 is involved in G-protein binding, while the N-terminal domain, homologous to other ADP-ribosylating toxins, contains the NAD(+)-binding site and the residues involved in catalysis. By site-directed mutagenesis and kinetic analyses Glu-129 and His-35 were identified as the catalytic residues. Glutamates analogous to Glu-129 are found in all studied ADP-ribosylating toxins, while His-35 is less well conserved. This suggests that Glu-129 acts on the common substrate NAD+, whereas His-35 plays its role on the acceptor substrates. We propose a mechanism in which Glu-129 exerts its action on the 2'-OH group of the NAD+ ribose, thereby facilitating the formation of an oxocarbonium-like intermediate and the weakening of the N-glycosidic bond. His-35 could increase the nucleophilicity of the cysteine in the G protein or the water molecule to attack the weakened N-glycosidic bond of NAD+ and yield the products of the reaction.
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PMID:A proposed mechanism of ADP-ribosylation catalyzed by the pertussis toxin S1 subunit. 852 86

BvgS and BvgA, a two-component system, regulate virulence gene expression in Bordetella pertussis. BvgS is a transmembrane sensor protein that can autophosphorylate and phosphorylate BvgA. Phosphorylated BvgA activates transcription of virulence genes. The cytoplasmic region of BvgS contains three domains separated by alanine/proline-rich sequences--the transmitter, receiver and C-terminus. We report that the C-terminal domain, like the transmitter and receiver, is an essential part of the phosphorelay from BvgS to BvgA. The BvgS C-terminal domain is phosphorylated in trans via a phosphotransfer mechanism by the cytoplasmic portion of BvgS, and trans-phosphorylation of the C-terminal domain requires both the transmitter and receiver. We also demonstrate that phosphorylated, purified C-terminal domain alone is sufficient for phosphotransfer to BvgA. A point mutation in the C-terminal domain (His1172-->Gln) abolishes BvgS activity in vivo and eliminates detectable phosphorylation of BvgA in vitro. Activity of BvgS His 1172-->Gln could be restored by providing the wild-type C-terminal domain in trans. Our results indicate an obligatory role for an alternate phosphodonor module in the BvgAS phosphorelay.
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PMID:Integration of multiple domains in a two-component sensor protein: the Bordetella pertussis BvgAS phosphorelay. 860 72

This work aimed to investigate the molecular role of gastrin in histamine synthesis in isolated rabbit fundic mucosal cells enriched in enterochromaffin-like (ECL) cells (37%). Gastrin stimulated histidine decarboxylase (HDC) activity by increasing the maximal velocity (Vmax) from 0.240 +/- 0.017 (basal value) to 0.332 +/- 0.012 pmol/mg protein/h and by decreasing the Michaelis-Menten constant value -Km; 73.90 +/- 2.2 vs. 93.42 +/- 4.32 microM (basal value)]. Pertussis toxin (PTX) (200 ng/ml) reduced the stimulation of HDC induced by 10 nM gastrin from 41.8 to 15.9%, whereas cholera toxin (CTX) (100 ng/ml) was without effect. Staurosporine and polymyxin B inhibited in a dose dependent manner the HDC activity stimulated by 10 nM gastrin. Phorbol 12-myristate 13-acetate (PMA; 100 nM) decreased Vmax (0.558 +/- 0.021 pmol/ mg protein/h) but did not change the Km. Furthermore, cycloheximide (0.1-10 microM) inhibited the gastrin-induced stimulation of HDC activity, whereas actinomycin D (up to 10 microM) was without effect. Finally, incubation of cells with gastrin (10 microM) left the expression of HDC mRNA unchanged. We concluded that gastrin, acting through "gastrin/CCK-B type" receptors coupled to PTX-sensitive G protein, exerts a short-term regulation of histamine synthesis in gastric ECL cells by increasing both the affinity of HDC for L-histidine and the number of active enzyme molecules. This last event, related to protein kinase C activation, could be due to a translational or posttranslational mechanism.
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PMID:Gastrin stimulation of histamine synthesis in enterochromaffin-like cells from rabbit fundic mucosa. 863 12

The unorthodox two-component sensor protein BvgS of Bordetella pertussis contains several interesting sequence motifs of unknown functional relevance, such as a histidine motif in its output domain, which is conserved among several unorthodox sensor proteins, a putative nucleotide binding site [Walker box type A] in its linker region, and a region in its periplasmic domain with significant homology to the TonB protein of Escherichia coli. We investigated potential functions of these sequences by constructing B. pertussis strains that express mutant BvgS derivatives. The His1172 residue in the output domain was exchanged for Gln, and the Walker motif was mutated either by the replacement of Lys625 by Arg, or of Gly624 by Val and Lys625 by Leu. To analyse the TonB motif, the periplasmic domain of BvgS was replaced with the corresponding domain of EvgS, an E. coli sensor that is highly homologous to BvgS but lacks the similarity with TonB. All mutations except the conservative Lys/Arg exchange in the Walker box caused the inactivation of BvgS, indicating the functional importance of the conserved motifs. The activity of the mutant proteins could be restored by complementation in trans with various separately expressed, truncated parts of BvgS. Mutations in the BvgS receiver domain could be complemented not only by a construct expressing the wild-type receiver and output domains, but also by the derivative containing the His-Gln exchange. Therefore, the histidine motif, although important for BvgS function, is not essential for complementation of BvgS mutants. The mutations in the Walker box and in the periplasmic domain could be complemented by a truncated BvgS derivative lacking the receiver and output domains. The characterization of a spontaneous revertant of the strain expressing the originally inactive EvgS/BvgS hybrid protein revealed the presence of a mutation in the BvgS linker region, conferring constitutive activity on the protein. As TonB energizes transport processes across the outer membrane of E. coli, the strain expressing the constitutive EvgS/BvgS hybrid protein lacking the TonB motif was used in preliminary investigations of a possible direct involvement of BvgS in transport processes.
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PMID:Conserved sequence motifs in the unorthodox BvgS two-component sensor protein of Bordetella pertussis. 880 90

Yellow fever, an acute mosquito-borne viral haemorrhagic fever, is preventable by use of the live, attenuated 17D vaccine. The vaccine is used principally in tropical climates and is subject to potentially adverse conditions. Lyophilized vaccine without stabilizers deteriorates rapidly when exposed to temperatures above -20 degrees C. In 1987, the WHO recommended that each lot of vaccine meet the following stability test: maintenance of potency (> 1,000 mouse i.c.LD50/human dose) with mean loss of titre < 1.0 log10 after being held at 37 degrees C for 14 days. In 1987, only 5 out of 12 yellow fever vaccines produced worldwide met the stability standards. To improve stability of the vaccine, a number of additives have been systematically investigated. A successful formulation, now used by a number of manufacturers, employs sugars, amino acids, and divalent cations [lactose (4%), sorbitol (2%), histidine (0.01 M), alanine (0.01 M), in phosphate buffered saline with Ca2+ and Mg2+]. As opposed to vaccine produced without stabilizers, which loses 1.5-2.5 log10/dose, stabilized vaccines lose only 0.3-0.5 log10 after being held at 37 degrees C for 14 days. The vaccine is stable after storage for > or = two years at 4 degrees C and 22 degrees C, and has a stability profile as good or better than many other live and inactivated vaccines currently used in the EPI, including measles, pertussis, oral and inactivated poliomyelitis vaccines. WHO is taking steps to enssure that all 11 current YF vaccine manufacturers produce vaccines that meet accepted stability standards. The principal rationale for increasing 17D vaccine stability beyond that achieved with the present stabilizers would be the improvement in stability of other EPI vaccines, to the point where yellow fever vaccine was the most sensitive vaccine among those deployed. The acceptable characteristics of current stabilized 17D vaccines and the high cost of changing and validating new vaccine formulations precludes a major investment at this time. Despite its stability when freeze dried, yellow fever 17D vaccine is quite unstable after reconstitution and must be discarded after one hour. Improvement in vaccine stability after reconstitution would thus reduce cost, stretch supplies of vaccine, and ensure against vaccine failures due to use of degraded vaccine. This is an area for future research.
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PMID:Stability of yellow fever vaccine. 885 20

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