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

The pSAM2 element of Streptomyces ambofaciens integrates site-specifically in the genome of different Streptomyces species by recombination between a 58 bp sequence common to the plasmid (attP) and the chromosome (attB). Southern hybridization analysis showed that sequences similar to the pSAM2 attB site were found in other actinomycetes (Mycobacterium, Nocardia, Micromonospora) as well as unrelated bacteria (Bacillus circulans, Escherichia coli, Clostridium botulinum, Bordetella pertussis, and Legionella pneumophila). Hybridizing fragments from B. circulans and Mycobacterium tuberculosis were cloned and sequenced. Comparison of these sequences with the sequence of the integration zone of S. ambofaciens revealed a conserved region of 76 bp which overlapped with the attB site. This conserved sequence was similar to the Salmonella typhimurium and E. coli tRNA(pro1) genes as well as a number of eucaryotic tRNA genes and had a proline-tRNA-like cloverleaf structure. Furthermore, the Streptomyces lividans attB site of the Streptomyces glaucescens element pIJ408 was also found to overlap a potential tRNA gene (tRNA(thr)). We note here that these two putative tRNA genes as well as those which overlap the attB site of the elements SLP1 of Streptomyces coelicolor and pMEA100 of Nocardia mediterranei all contain the site where integrative recombination takes place. These presumptive actinomycete tRNA genes lack the 3' terminal CCA sequence found in most procaryotic tRNA genes.
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PMID:The chromosomal integration site of the Streptomyces element pSAM2 overlaps a putative tRNA gene conserved among actinomycetes. 170 70

The S1 subunit (Mr 28,000) of pertussis toxin expresses thiol-dependent enzymatic ADP-ribosyltransferase and NAD-glycohydrolase activities. Site-directed mutagenesis experiments were performed on the codon for Cys-41 of this subunit to investigate the role of this residue in both enzymatic activities. Deletion of Cys-41 caused a decrease in both activities below detectable levels, whereas replacement of this residue by serine, glycine, proline, or asparagine only slightly reduced the activities. The enzymatic activities of these mutants were thiol-independent. The deletion of Ser-40, adjacent to Cys-41, again caused reduction of the enzymatic activities to undetectable levels. Steady-state kinetic experiments showed that the kcat of the mutant protein in which Cys-41 was replaced by glycine was nearly identical to the kcat of the parent version. However, the Km for NAD of the mutant was significantly higher relative to that of the wild type version. These results indicate that the side-chain of Cys-41 is not essential for enzymatic activities and that Cys-41 is not involved in the rate of catalysis but is probably located at or close to the NAD-binding site. The introduction of a negative charge at position 41 through the replacement of Cys-41 by either aspartate or glutamate reduced the enzymatic activities to very low but measurable levels, suggesting a charge-charge repulsive interaction between these residues and possibly one or both of the phosphates of NAD. Cys-41 may therefore be located close to the phosphate subsite of the NAD-binding site.
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PMID:The role of cysteine 41 in the enzymatic activities of the pertussis toxin S1 subunit as investigated by site-directed mutagenesis. 215 32

We tested the effects that pertussis toxin had on bone resorption mediated by cAMP-dependent and cAMP-independent stimuli in 19-day-old fetal rat long bones. Agents that stimulate cAMP were PTH, prostaglandin E2, and calcitonin. Agents that act independent of cAMP were: phorbol 13-myristate 12-acetate (PMA), 1,25-dihydroxyvitamin D3, murine interleukin-1 alpha, osteoclast-activating factor, and human tumor necrosis factor-alpha. Pertussis (1-10 ng/ml) produced a dose-related inhibition of resorption in unstimulated control cultures. The inhibitory effect was not associated with changes in either [3H]thymidine or [3H]proline incorporation into bones. beta-Glucuronidase activity in the medium was decreased. PMA was the only agonist whose resorptive effect was completely blocked by pertussis. The resorptive response to other stimulators was reduced, but treated/control ratios usually remained the same or increased because of the greater effect of pertussis on control resorption. There was a partial inhibition of the resorptive effect of low doses of prostaglandin E2 (10 nM), but increasing the concentration of agonist overcame the inhibition. Pertussis did not enhance the sensitivity of bones to calcitonin. Pertussis enhanced the cAMP response to PTH, but had no effect on basal cAMP production. Since PMA was inhibited by pertussis while agents that may act through cAMP-mediated or phosphatidylinositol pathways were not affected, we hypothesize that a protein kinase-C dependent pathway can modulate bone resorption.
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PMID:Effects of pertussis toxin on resorption of 19-day-old fetal rat long bones. 253 69

The amino acid consumption by Bordetella pertussis growing in broth containing casein hydrolysate was examined. Serine, proline, alanine, glycine, aspartate, and glutamate were rapidly consumed, in a manner which suggested that they supplied the energy requirements of the organism; exhaustion of the energy source appeared to be the main factor limiting the yield of cells. There was no correlation between the utilization of individual amino acids and the phase of growth; uptake appeared to depend only upon relative concentrations. Consumption of threonine, phenylalanine, histidine, leucine, and methionine was slight; consumption of valine and lysine was variable, and isoleucine was excreted. The addition of monosodium l-glutamate (3 mg/ml) to the broth in shaken flasks increased the cell yield by an average of 43.5%. It had no detectable adverse effect upon the agglutin-producing capacity, agglutinability in antisera versus smooth and rough growth phases, mouse-lethal toxicity, histamine-sensitizing factor potency, or intracerebral protective potency of the culture. Broth supplemented with monosodium l-glutamate has been used over a 2-year period to prepare experimental vaccines by both batch and continuous cultivation methods at controlled pH; the cell yields obtained from the supplemented broth have been up to 52% higher than those from the basal broth. The use of glutamate to replace a proportion of casein hydrolysate in the broth caused a reduction in the cell yield, an alteration in cell morphology, and reduction in the mouse-lethal toxicity, the histamine-sensitizing factor potency, and the intracerebral protective potency of the cells.
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PMID:Use of glutamic acid to supplement fluid medium for cultivation of Bordetella pertussis. 431 42

We report here the identification of a virulence-associated factor, Tcf, (tracheal colonization factor), produced by strains of Bordetella pertussis but not Bordetella parapertussis or Bordetella bronchiseptica. This protein is encoded by the tcfA gene. When a strain of B. pertussis 18323 lacking this protein is used to infect mice with an aerosol challenge, the number of bacteria isolated from the tracheas is decreased 10-fold when compared with the parent 18323. The derived amino acid sequence of tcfA predicts a 68 kDa RGD-containing, proline-rich protein, which after cleavage of a typical prokaryotic signal sequence would be 64 kDa. Amino acid sequence analysis demonstrates that the C-terminal 30 kDa of this protein shows 50% identity to the 30 kDa C-terminus of another Bordetella protein, the pertactin precursor. The N-terminal 34 kDa region contains the three amino-acid motif RGD and is 16.5% proline. Coupled in vitro transcription and translation analysis indicates that the tcfA gene product migrates as two bands of approximately 90 kDa. A fusion protein of the N-terminal, 34 kDa portion of Tcf to maltose-binding protein migrates, on SDS-PAGE, 30 kDa higher than expected from the combined molecular weights. Polyclonal antisera raised against the unique N-terminal portion of Tcf recognizes 90 kDa and 60 kDa bands in immunoblots of whole-cell lysates of strains of B. pertussis; it does not recognize any protein in whole-cell lysates of B. bronchiseptica or B. parapertussis. Supernatants of cultures of B. pertussis 18323 contain the 60 kDa form of the protein. Southern blot analysis of chromosomal DNA from strains of B. bronchiseptica and B. parapertussis, using a probe derived from tcfA, shows strong hybridization only to B. pertussis DNA. Thus, Tcf appears to be a unique virulence factor of B. pertussis.
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PMID:Tracheal colonization factor: a Bordetella pertussis secreted virulence determinant. 747 58

Antibody-binding domains on the major subunits of Bordetella pertussis serotype 2 (Fim2) and 3 fimbriae (Fim3) have been identified using synthetic peptides which were screened for recognition by anti-protein monoclonal antibodies (mAbs). The presence of non-contiguous fimbrial epitopes was demonstrated by both anti-Fim2 and anti-Fim3 mAbs, several of which recognized at least two peptides that were discontinuous in the amino acid sequence of the corresponding subunits. The specificity of one mAb, 51/24, directed against Fim2, was investigated by replacement-set analysis of a 10-residue peptide, and revealed that antibody binding to the peptide was dependent on the sequence N94PQ96 which is non-conserved in Fim3. Furthermore, proline at residue 95 was found to be essential for mAb 51/24 binding. The specific anti-Fim3 mAb, AG3A, was found to recognize the 10-residue carboxy-terminal peptide from both Fim3 and, unexpectedly, from Fim2. This result suggests that mAb AG3A serospecificity at the protein level is determined by a conformational constraint which prevents mAb AG3A binding to the Fim2 C-terminal domain. Several free peptides containing amino acid residues which comprise part of the Fim2 and Fim3 epitopic domains were prepared as immunogens. One of these peptides was immunogenic in the mouse, indicating the location of a T-helper cell epitope within the peptide sequence, and induced a strong anti-peptide antibody response. The other peptides each required immunization as a conjugate with a carrier protein for anti-peptide antibody stimulation. All four anti-peptide antibody preparations only weakly recognized fimbriae-coated ELISA plates.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Localization of antigenic domains on the major subunits of Bordetella pertussis serotype 2 and 3 fimbriae. 751 70

ADP-ribosylating protein exotoxins from Vibrio cholerae (CT) and Escherichia coli (LT-I) share two short regions of sequence similarity with Bordetella pertussis toxin (PT). Previous studies have indicated that substitution of arginine for lysine 7 within the first region of CT drastically decreases ADP ribosyltransferase activity. We have more closely defined the role of other amino acids in this region by generating modified proteins in which arginine 7 was replaced with lysine (R7K), aspartate 9 was replaced with arginine (D9R), glycine was substituted for proline 12 (P12G), amino acids 6 to 13 were deleted (delta 613) or the C-terminal KDEL sequence was changed to NEDL. The modified proteins R7K, D9R and delta 613 exhibited undetectable ADP ribosyltransferase activity. Comparison of the tryptic digest of R7K with native CT suggested that changes in protein conformation may be responsible for the loss of ADP-ribosylation activity.
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PMID:Expression and mutagenesis of recombinant cholera toxin A subunit. 772 60

The proline-arginine (PR) -rich antibacterial peptide, PR-39, kills bacteria by a non-pore-forming mechanism. Because this neutrophil peptide possesses several distinct functional properties and because other antimicrobial peptides are chemoattractants, we sought to determine whether PR-39 was a chemoattractant for porcine leukocytes. The peptide was synthesized by the solid-phase method using t-Boc chemistry and purified by reversed-phase high-performance liquid chromatography. Leukocyte migration was assessed with the use of a 48-well microchemotaxis chamber. PR-39 induced the directed migration of neutrophils. The peak chemotaxis response occurred at 0.5-2 microM, which was slightly lower than the minimal inhibitory and bactericidal concentrations for PR-39. However, the peptide was not a chemoattractant for mononuclear cells. Truncation of PR-39 suggested that the neutrophil chemoattractant domain may be contained within the first 26 amino acid residues. Intracellular Ca2+ fluxes in response to PR-39 were monitored by flow cytometry and showed a transient increase that peaked at approximately 40 s and approached basal values by 4 min. However, in a Ca2+-free environment, the PR-39-induced Ca2+ increase was abrogated. Furthermore, PR-39 did not induce neutrophil chemotaxis in the absence of extracellular Ca2+, and pertussis toxin inhibited both neutrophil chemotaxis and Ca2+ mobilization. Taken together, these data suggest that PR-39 is a Ca2+-dependent chemoattractant of neutrophils. The finding of a neutrophil antibacterial peptide that is also a neutrophil chemoattractant is intriguing and may indicate an important role for PR-39 in inflammation.
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PMID:Chemoattractant properties of PR-39, a neutrophil antibacterial peptide. 912 12

Cysteine351 is the site for pertussis toxin-catalyzed ADP-ribosylation in the G protein Gi1 alpha. Alteration of this residue, or the equivalent cysteine in other Gi-family G proteins, has been used to examine specific interactions between receptors and these G proteins. However, no systematic analysis has been performed to determine the quantitative effect of such alterations. To address this we mutated cysteine351 of Gi1 alpha to all other possible amino acids. Each of the G protein mutants was transiently coexpressed along with the porcine alpha 2A-adrenoceptor in HEK 293/T cells. Following pertussis toxin treatment of the cells, membranes were prepared and the capacity of the agonist UK14304 to stimulate the binding of [35S]GTP gamma S to the modified G proteins was measured. A spectrum of function was observed. The presence of either a charged amino acid or a proline at this position essentially attenuated agonist regulation. The wild-type G protein did not result in maximal stimulation by agonist. The presence of certain branched chain aliphatic amino acids or bulky aromatic R groups at amino acid351 resulted in substantially greater maximal stimulation by the alpha 2A-adrenoceptor than that achieved with the wild-type sequence. The degree of activation of the forms of Gi1 alpha correlated strongly with the octanol/water partition coefficient of the amino acid at residue351. Variation in EC50 values for agonist-induced stimulation of binding of [35S]GTP gamma S to the mutant G proteins also correlated with the octanol/water partition coefficient. These results define a central role for hydrophobicity of this residue in defining productive receptor-G protein interactions.
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PMID:Hydrophobicity of residue351 of the G protein Gi1 alpha determines the extent of activation by the alpha 2A-adrenoceptor. 970 91

The recently identified 17-amino acid peptide nociceptin (orphanin FQ) is the endogenous ligand for the opioid receptor-like-1 (ORL-1) receptor. A physiologic role for nociceptin (OFQ) activation of the ORL-1 receptor (OFQR) may be to modulate opioid-induced analgesia. The molecular mechanism by which nociceptin (OFQ) and ORL-1 (OFQR) modify opioid-stimulated effects, however, is unclear. Both ORL-1 (OFQR) and opioid receptors mediate pertussis toxin (PTX)-sensitive signal transduction, indicating these receptors are capable of coupling to Gi/Go proteins. This study determines that nociceptin stimulates an intracellular signaling pathway, leading to activation of mitogen-activated protein (MAP) kinase in CHO cells expressing ORL-1 receptor (OFQR). Nociceptin (OFQ)-stimulated MAP kinase activation was inhibited by PTX or by expression of the carboxyl terminus of beta-adrenergic receptor kinase (betaARKct), which specifically blocks Gbetagamma-mediated signaling. Expression of the proline-rich domain of SOS (SOS-PRO), which inhibits SOS interaction with p21ras, also attenuated nociceptin (OFQ)-stimulated MAP kinase activation. The phosphatidylinositol 3-kinase (PI-3K) inhibitors wortmannin and LY294002 reduced nociceptin (OFQ)-stimulated MAP kinase activation, whereas inhibition of protein kinase C (PKC) activity by bisindolylmaleimide I or cellular depletion of PKC had no effect. In a similar manner, in cells expressing mu-opioid receptor, [D-Ala2,N-Me-Phe4,Gly-ol]-enkephalin (DAMGO; a mu-opioid receptor-selective agonist) stimulated PTX-sensitive MAP kinase activation that was inhibited by wortmannin, LY294002, betaARKct expression, or SOS-PRO expression but not affected by inhibition of PKC activity. These results indicate that both ORL-1 (OFQR) and mu-opioid receptors mediate MAP kinase activation via a signaling pathway using the betagamma-subunit of Gi, a PI-3K, and SOS, independent of PKC activity. In cells expressing both ORL-1 (OFQR) and mu-opioid receptors, pretreatment with nociceptin decreased subsequent nociceptin (OFQ)- or DAMGO-stimulated MAP kinase activation. In contrast, pretreatment of cells with DAMGO decreased subsequent DAMGO-stimulated MAP kinase but had no effect on subsequent nociceptin (OFQ)-stimulated MAP kinase activation. These results demonstrate that nociceptin (OFQ) activation of ORL-1 (OFQR) can modulate mu-opioid receptor signaling in a cellular system.
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PMID:Nociceptin (ORL-1) and mu-opioid receptors mediate mitogen-activated protein kinase activation in CHO cells through a Gi-coupled signaling pathway: evidence for distinct mechanisms of agonist-mediated desensitization. 972 27


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