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Pertussis Toxin fermentation

I Kaji, Y Akiba, T Furuyama, D W Adelson, K Iwamoto, M Watanabe, A Kuwahara, J D Kaunitz
BACKGROUND: Short-chain fatty acids (SCFA) are microbial fermentation products absorbed by the colon. We recently reported that activation of the SCFA receptor termed free fatty acid receptor 3 (FFA3), expressed on cholinergic nerves, suppresses nicotinic acetylcholine receptor (nAChR)-mediated transepithelial anion secretion. This study aimed to clarify how activation of neurally expressed FFA3 affects colonic motor function. METHODS: FFA3-expressing myenteric neurons were identified by immunostaining; contractions of isolated circular muscle strips obtained from rat proximal colon were measured by isometric transducers...
January 2018: Neurogastroenterology and Motility: the Official Journal of the European Gastrointestinal Motility Society
Carolina Iraporda, Agustina Errea, David E Romanin, Delphine Cayet, Elba Pereyra, Omar Pignataro, Jean Claude Sirard, Graciela L Garrote, Analía G Abraham, Martín Rumbo
The use of short chain fatty acids to modulate gastrointestinal inflammatory conditions such as ulcerative colitis has produced encouraging results either in animal models or also in clinical trials. Identifying the key cellular and molecular targets of this activity will contribute to establish the appropriate combinations/targeting strategies to maximize the efficacy of anti-inflammatory interventions. In the present work, we evaluated in vitro the interaction of lactate, acetate, propionate and butyrate on cells relevant for innate immune response of the gastrointestinal tract...
October 2015: Immunobiology
Elen A Perpetuo, Ivo Lebrun, Luis Juliano, Maria Aparecida Juliano, Maria Aparecida Sakauchi, Sally M A Prado
Proteases were identified and characterized from the culture supernatant of the C. diphtheriae and B. pertussis bacteria. The proteases were secreted in the media and detected at the end of the exponential growth phase. Activity was detected in some fluorescent substrates, based on selected protein sequences such as insuline beta-chain, bradykinin, and synaptobrevin. The proteases were purified by means of gel filtration chromatography. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of the purified proteins indicated, for the main secreted proteins, an estimated molecular mass of 30 kDa in C...
2007: Preparative Biochemistry & Biotechnology
J A Bogdan, J Nazario-Larrieu, J Sarwar, P Alexander, M S Blake
Pertussis toxin (Ptx) expression and secretion in Bordetella pertussis are regulated by a two-component signal transduction system encoded by the bvg regulatory locus. However, it is not known whether the metabolic pathways and growth state of the bacterium influence synthesis and secretion of Ptx and other virulence factors. We have observed a reduction in the concentration of Ptx per optical density unit midway in fermentation. Studies were conducted to identify possible factors causing this reduction and to develop culture conditions that optimize Ptx expression...
November 2001: Infection and Immunity
L V Miriasova, I A Basnak'ian, T V Kopaeva, I Iu Sarkisov
The processes of the cultivation of Bordetella pertussis, immobilized on polyurethane carrier in a fermenter, were carried out and studied. Acellular pertussis preparations were produced from the culture fluid obtained in the batch and multi-cycle cultivation processes with immobilized cells, as well as in the process with interrupted fermentation (for confirming the possibility of the preservation of cell viability). The content of protein and B. pertussis toxin in these preparations, as well as their leukocytes-stimulating and hemagglutinating activity, did not differ from similar characteristics of preparations obtained from culture fluid in homogeneous cultivation...
May 1999: Zhurnal Mikrobiologii, Epidemiologii, i Immunobiologii
P Chong, M Klein
A general procedure for purifying biologically active pertussis toxin from Bordetella pertussis fermentation broth using affinity chromatography on heat-treated fetuin-Sepharose CL-4B is described. Diethanolamine is used as eluent in this single-step purification to prepare endotoxin-free pertussis toxin in good yield (70%) and high purity (greater than 95%). This one-step affinity chromatography procedure can be easily applied for large-scale preparation of pertussis toxin S1 subunit and its B-component. The affinity-purified S1 subunit is devoid of any of the histamine-sensitizing activity normally associated with pertussis toxin...
July 1989: Biochemistry and Cell Biology, Biochimie et Biologie Cellulaire
N Andorn, J B Kaufman, T R Clem, R Fass, J Shiloach
No abstract text is available yet for this article.
1990: Annals of the New York Academy of Sciences
P Chong, G Jackson, W Cwyk, M Klein
A simple and rapid method for the simultaneous determination of Bordetella pertussis toxin (PT) and filamentous haemagglutinin (FHA) concentrations in fermentation broths has been developed. The rapid single-step analysis performed by hydroxyapatite high-performance liquid chromatography using a salt gradient with UV detection allows both the separation of PT from FHA and the measurement of their respective concentrations. The assay is highly reproducible. Over 35 lots of acellular B. pertussis vaccine production lots were examined and PT concentrations measured by high-performance liquid chromatography were found to be in good agreement with the values obtained by sodium dodecyl sulphate-polyacrylamide gel electrophoresis densitometry...
July 20, 1990: Journal of Chromatography
M D Oxer, C M Bentley, J G Doyle, T C Peakman, I G Charles, A J Makoff
The anaerobically-regulated nirB promoter was used to express heterologous genes in Escherichia coli. Under anaerobic conditions the promoter was able to express tetanus toxin fragment C at approximately 20% total cell protein (tcp) and the Bordetella pertussis antigen pertactin at greater than 30% tcp. These levels are comparable to those obtained for the same products using the tac promoter. The nirB promoter is very well regulated, giving almost two orders of magnitude increase in fragment C on complete removal of oxygen...
June 11, 1991: Nucleic Acids Research
G R Zealey, S M Loosmore, R K Yacoob, S A Cockle, A B Herbert, L D Miller, N J Mackay, M H Klein
Nontoxic analogs of pertussis toxin (PT), produced by in vitro mutagenesis of the tox operon, are immunogenic and protective against infection by Bordetella pertussis. The moderate levels of PT production by B. pertussis, however, make it the limiting antigen in the formulation of multicomponent, acellular, recombinant whooping cough vaccines. To increase production of the highly detoxified Lys9Gly129 PT analog by B. pertussis, additional copies of the mutated tox operon were integrated into the bacterial chromosome at the tox or fha locus by unmarked allelic exchange...
January 1992: Applied and Environmental Microbiology
G R Zealey, S M Loosmore, R K Yacoob, S A Cockle, L J Boux, L D Miller, M H Klein
We replaced the wild-type TOX operon of Bordetella pertussis with in vitro mutated, detoxified alleles by electroporetic transformation using unmarked linear DNA. Uptake of DNA was selected by transient ampicillin resistance and two simultaneous recombination events resulted in gene-replacement at the natural locus with no integration of heterologous DNA. TOX alleles were stable without selection and recombinant strains secreted non-toxic, fully assembled, protective pertussis toxin (PT) analogues with kinetics similar to the parental vaccine strain under production-scale fermentation conditions...
November 1990: Bio/technology
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