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)

A truncated Bordetella pertussis cya gene product was expressed in Escherichia coli and purified by affinity chromatography on calmodulin-agarose. Trypsin cleavage of the 432-residue recombinant protein (Mr = 46,659) generated two fragments of 28 kDa and 19 kDa. These fragments, each containing a single Trp residue, were purified and analyzed for their catalytic and calmodulin-binding properties. The 28-kDa peptide, corresponding to the N-terminal domain of the recombinant adenylate cyclase, exhibited very low catalytic activity, and was still able to bind calmodulin weakly, as evidenced by using a fluorescent derivative of the activator protein. The 19-kDa peptide, corresponding to the C-terminal domain of the recombinant adenylate cyclase, interacted only with calmodulin as indicated by a shift in its intrinsic fluorescence emission spectrum or by the enhancement of fluorescence of dansyl-calmodulin. T28 and T19 fragments exhibited an increased sensitivity to denaturation by urea as compared to uncleaved adenylate cyclase, suggesting that interactive contacts between ordered portions of T28 and T19 in the intact protein participate both in their own stabilization and in stabilization of the whole tertiary structure. The two fragments reassociated into a highly active calmodulin-dependent species. Reassociation was enhanced by calmodulin itself, which 'trapped' the two complementary peptides into a stable, native-like, ternary complex, which shows similar catalytic properties to intact adenylate cyclase.
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PMID:Isolation and characterization of catalytic and calmodulin-binding domains of Bordetella pertussis adenylate cyclase. 200 7

degP-deficient strains of Escherichia coli grown in M-9 medium supplemented with ZnCl2 expressed the recombinant S1 subunit of pertussis toxin (rS1) in a form electrophoretically identical to the authentic S1 subunit. Subcellular fractionation showed that the full-length form of rS1 was membrane associated, while proteolytic fragments of rS1 were present in the periplasm. rS1 was extracted from outer membrane preparations with 8 M urea and purified by gel filtration chromatography. Purified rS1 ADP-ribosylated transducin at a similar molar efficiency relative to authentic pertussis toxin and, when associated with the native B oligomer of pertussis toxin, elicited Chinese hamster ovary cell clustering.
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PMID:Biochemical and biological activities of recombinant S1 subunit of pertussis toxin. 210 94

Initially we established that the binding of collagen to human blood platelets stimulates both the rapid loss of PIP2 and the generation of inositol-4,5-bisphosphate (IP2) and inositol-1,4,5-triphosphate (IP3). These results indicate that the binding of collagen stimulates inositol phospholipid-specific phospholipase C during platelet activation. The fact that GTP or GTP-gamma-S augments, and pertussis toxin inhibits, collagen-induced IP3 formation suggests that a GTP-binding protein (or (or proteins) may be directly involved in the regulation of phospholipase C-mediated phosphoinositide turnover in human platelets. We have used several complementary techniques to isolate and characterize a platelet 41-kDa polypeptide (or polypeptides) that has a number of structural and functional similarities to the regulatory alpha i subunit of the GTP-binding proteins isolated from bovine brain. This 41-kDa polypeptide (or polypeptides) is found to be closely associated with at least four membrane glycoproteins (e.g., gp180, gp110, gp95, and gp75) in a 330-kDa complex that can be dissociated by treatment with high salt plus urea. Most important, we have demonstrated that antilymphoma 41-kDa (alpha i subunit of GTP-binding proteins) antibody cross-reacts with the platelet 41-kDa protein (or proteins) and the alpha i subunit of bovine brain Gi alpha proteins, and blocks GTP/collagen-induced IP3 formation. These data provide strong evidence that the 41-kDa platelet GTP-binding protein (or proteins) is directly involved in collagen-induced signal transduction during platelet activation.
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PMID:Membrane-associated 41-kDa GTP-binding protein in collagen-induced platelet activation. 211 41

Three distinct antipeptide antisera generated against synthetic peptides that represent parts of the primary sequence of the alpha-subunit of the (pertussis toxin-sensitive) guanine nucleotide binding protein G0 were used in two-dimensional immunoblots of membranes of neuroblastoma X glioma (NG108-15) cells. Each antiserum identified two distinct polypeptides of some 39 kDa. These had apparent isoelectric points of 5.5 and 5.8. Differentiation of NG108-15 cells in response separately to dibutyryl cyclic AMP (cAMP), 8-bromo cAMP, forskolin, and prostaglandin E1 produced elevated levels of G0 alpha, as has previously been noted in one-dimensional immunoblots. Two-dimensional analysis demonstrated that the cAMP-induced increases in levels of G0 alpha were only of the more acidic isoform. The two isoforms were both substrates for pertussis toxin-catalysed ADP-ribosylation and did not appear to represent differentially phosphorylated forms of the same polypeptide. Separation of the two forms of G0 alpha could be achieved in one-dimensional sodium dodecyl sulphate-polyacrylamide gel electrophoresis when 4 M deionized urea was included in the resolving gel. The more slowly migrating band was the acidic form and corresponded exactly in mobility with the major form of G0 from both rat and mouse brain. There was no equivalent in brain of the more rapidly migrating form of G0 from the cells. In agreement with the data from two-dimensional gels, only the more slowly migrating form was expressed in considerably higher amounts following cAMP-induced differentiation of NG108-15 cells. Of these two forms of "G0," the acidic species is equivalent to G0 from brain, but the basic form is not identical with G0*, which has been purified from bovine brain.
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PMID:Identification of two distinct isoforms of the guanine nucleotide binding protein G0 in neuroblastoma X glioma hybrid cells: independent regulation during cyclic AMP-induced differentiation. 217 64

Bordetella pertussis, the pathogen responsible for whooping cough, produces a calmodulin-sensitive adenylate cyclase. Several investigators have shown that the partially purified adenylate cyclase is capable of entering animal cells and elevating intracellular cAMP levels (Confer and Eaton: Science 217:948-950, 1982; Shattuck and Storm: Biochemistry 24:6323-6328, 1985). However, the mechanism for entry of the catalytic subunit of this adenylate cyclase into animal cells is unknown. It has been reported that the B. pertussis adenylate cyclase extracted from bacterial cells with urea does not enter animal cells by receptor-mediated endocytosis. There is, in addition to the cell associated form of the B. pertussis adenylate cyclase, a cell-invasive form of the enzyme secreted into the bacterial culture media. The properties of the cell-associated and secreted enzymes are significantly different (Masure and Storm: Biochemistry 28:438-442, 1989). In this study, we report evidence that the secreted form of the B. pertussis adenylate cyclase enters animal cells by a mechanism distinct from receptor-mediated endocytosis.
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PMID:Evidence that the adenylate cyclase secreted from Bordetella pertussis does not enter animal cells by receptor-mediated endocytosis. 217 58

Bordetella pertussis produces a number of virulence determinants which contribute to its pathogenicity. One factor, the adenylate cyclase toxin (ACT), has been suggested to directly penetrate human phagocytes and disrupt their normal function by direct production of intracellular cyclic AMP (cAMP). Experiments evaluating the production of cell-associated ACT in liquid cultures of B. pertussis 504 demonstrated that the greatest activity was observed during mid-log-phase growth. Urea extracts of cells harvested during the time of maximal ACT production have been used to purify the toxin with both biological and enzymatic activities. ACT is a protein with an apparent molecular mass of 220 kDa and an isoelectric point of 7.0. The specific activity of purified ACT is 17,000 mumol of cAMP formed per mg per min. The the biological specific activity of purified ACT is 6,250 nmol of intracellular cAMP formed per mg per min in 2 x 10(6) S49 lymphoma cells per ml. Preparations containing 8 micrograms of ACT completely abrogated the chemiluminescence response of 2 x 10(6) human neutrophils per ml.
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PMID:Adenylate cyclase toxin of Bordetella pertussis: production, purification, and partial characterization. 222 32

A truncated, 432 residue long, Bordetella pertussis adenylate cyclase expressed in Escherichia coli was analyzed for intrinsic fluorescence properties. The two tryptophans (Trp69 and Trp242) of adenylate cyclase, each situated in close proximity to residues important for catalysis or binding of calmodulin (CaM), produced overlapping fluorescence emission bands upon excitation at 295 nm. CaM, alone or in association with low concentrations of urea, induced important modifications in the spectra of adenylate cyclase such as shifts of the maxima and change in the shape of the bands. From these changes and from the fluorescence spectrum of a modified form of adenylate cyclase, in which a valine residue was substituted for Trp242, it was deduced that, upon binding of CaM to the wild-type adenylate cyclase, only the environment of Trp242 was affected. The fluorescence maximum of this residue, which is more exposed to the solvent than Trp69 in the absence of CaM, is shifted by 13 nm to shorter wavelength upon interaction of protein with its activator. Trypsin cleaved adenylate cyclase into two fragments, one carrying the catalytic domain, and the second carrying the CaM-binding domain (Ladant et al., 1989). The isolated peptides conserved most of the environment around their single tryptophan residues, as in the intact adenylate cyclase, which suggests that the two domains of truncated B. pertussis adenylate cyclase also conserved most of their three-dimensional structure in the isolated forms.
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PMID:Intrinsic fluorescence of a truncated Bordetella pertussis adenylate cyclase expressed in Escherichia coli. 226 68

The mechanism of heterologous desensitization of adenylate cyclase stimulation was studied in cultured neonatal rat heart muscle cells. After culturing of the cells for 3 days in the presence of 1 microM noradrenaline there was in addition to a 52% decrease in isoproterenol-stimulated adenylate cyclase activity, a lessening of the stimulation of beta-adrenoceptor-independent adenylate cyclase by guanosine-5'-O-(thiotriphosphate) and forskolin by 24 and 34%, respectively. The decrease in receptor-independent adenylate cyclase stimulation by forskolin, but not the attenuation of isoproterenol-stimulated adenylate cyclase activity, was abolished by pertussis toxin (PTX) pretreatment of the cells. Gi, the inhibitory G-protein of adenylate cyclase was therefore quantitated. Labelling of the Mr approximately 40 kDa PTX substrates in membranes of noradrenaline-treated cells was increased by 70% as shown by pertussis toxin-catalyzed ADP ribosylation of heart cell membranes. This increase was also seen in the presence of an excess of purified beta gamma-subunits of transducin and of GTP, suggesting that the increased labelling was not due to elevation of the level of beta gamma-subunits or increase in the concentration of GTP in the membranes of noradrenaline-treated cells. Analysis of the PTX substrates on high resolution urea/SDS-polyacrylamide gels revealed that at least two distinct PTX substrates (40 and 41 kDa) were present in rat heart cell membranes. The labelling of both substrates was increased in membranes of desensitized cells. Immunoblotting of heart cell membranes with anti-Gi alpha-antibodies demonstrated a marked increase in the amount of Gi alpha in membranes of noradrenaline-treated cells. In contrast, immunoblotting with anti-beta-antibodies showed that the level of the beta-subunit of G-proteins (36 kDa) was unchanged after noradrenaline exposure. The data indicate that prolonged treatment of rat heart muscle cells with noradrenaline leads to an increase in the level of alpha-subunits of Gi-proteins. This suggests that this increase is responsible for the observed heterologous desensitization of adenylate cyclase stimulation.
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PMID:Mechanism of noradrenaline-induced heterologous desensitization of adenylate cyclase stimulation in rat heart muscle cells: increase in the level of inhibitory G-protein alpha-subunits. 250 67

Bordetella pertussis organisms secrete adenylate cyclase, at least one form of which can invade host cells and appears to be a virulence factor. Treatment of urea extracts containing invasive cyclase of B. pertussis with trypsin, chymotrypsin, or subtilisin abolishes the ability to increase intracellular cyclic AMP levels in CHO cells (invasiveness) at concentrations that have minimal or no effects on adenylate cyclase activity. Higher protease concentrations can inhibit catalytic activity, and 1 microM calmodulin protects this catalytic activity, but not invasiveness, against proteolytic inhibition. Rabbit immunoglobulin G (IgG) fractions from antisera prepared against urea extracts inhibited invasiveness at 10-fold-lower concentrations than inhibited catalytic activity. One IgG from a rabbit immunized against a partially purified, noninvasive form of the B. pertussis adenylate cyclase inhibited catalytic activity but was ineffective against invasiveness. We conclude that these two properties of the adenylate cyclase are independent functions that reside on different domains of the same protein or on different proteins.
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PMID:Dissociation of catalytic and invasive activities of Bordetella pertussis adenylate cyclase. 254 62

A study was conducted to investigate nephritogenic tubular basement membrane antigens common to human and rat kidneys. Brown Norway (BN) rats were immunized with human renal basement membrane in complete Freund's adjuvant simultaneously with Bordetella pertussis vaccine. The immunized rats developed polyuria and increased levels of serum creatinine one week after the second immunization. Renal histology at this time revealed marked, acute tubulointerstitial nephritis with linear deposition of IgG and C3 along the tubular basement membrane and Bowman's capsule, but not along the glomerular basement membrane. Rats with this tubulointerstitial nephritis rapidly developed antibodies against renal antigens from normal BN rats such as tubular basement membrane and proximal tubule brush border, however antibodies to glomerular basement membrane appeared later. Western blotting using the same rat sera detected a 145-kDa antigen from 8 M urea-solubilized human renal basement membrane and 120-kDa, 135-kDa and 145-kDa antigens from 8 M urea-solubilized BN rat renal basement membrane. This suggests that renal basement membranes of human and rat origin have common antigens involved in the pathogenesis of tubulointerstitial nephritis.
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PMID:Induction of interstitial nephritis in rats by basement membrane of human origin. 268 56


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