Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.4 (trypsin)
 42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A new pro-carboxypeptidase (pCPB) gene was cloned by RT-PCR from SD rat pancreas and its overexpression in Escherichia coli resulted in the formation of inclusion bodies (IBs). The IBs of pCPB were solubilized in 8 M urea and successively refolded by urea gradient gel filtration. Subsequently, the renatured pCPB was digested by trypsin. Recombinant active CPB was obtained by passing through DEAE-FF ion exchange and Sephadex-G100 chromatographic column. Capillary electrophoresis assay showed that the purity of the recombinant CPB (rCPB) exceeded 90%. Further, some properties of rCPB were characterized. The optimum of activity was achieved at pH 7-9. The activity of rCPB was inhibited by typical metal chelating agents (EDTA) and Hg2+, and was activated by Co2+ and heat treatment at 40 degrees C. The two-dimension electrophoresis map of rCPB showed that the pI value of rCPB was 5.35. UV absorbance spectrum of the enzyme showed that an absorbance maximum was at 277 nm.
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PMID:The renaturation of procarboxypeptidase B by urea gradient gel filtration and some properties of recombinant carboxypeptidase B. 1652 83

Clostridium perfringens is capable of producing up to 15 toxins, including alpha-toxin (CPA), beta-toxin (CPB), epsilon-toxin (ETX), enterotoxin, beta2-toxin (CPB2), and perfringolysin O. Type B isolates, which must produce CPA, CPB, and ETX, are associated with animal illnesses characterized by sudden death or acute neurological signs, with or without intestinal damage. Type B pathogenesis in ruminants is poorly understood, with some animals showing lesions and clinical signs similar to those caused by either type C or type D infections. It is unknown whether host or environmental conditions are dominant for determining the outcome of type B disease or if disease outcomes are determined by variable characteristics of type B isolates. To help clarify this issue, 19 type B isolates were evaluated for toxin production during late-log-phase growth via quantitative Western blotting and by biological activity assays. Most type B isolates produced CPB levels similar to those produced by type C isolates in vitro and have the potential to produce genotype C-like disease. The lethality of type B isolate supernatants administered intravenously to mice was evaluated with or without prior trypsin treatment, and monoclonal antibody neutralization studies also were performed. Correlation analyses comparing toxin levels in type B supernatants versus lethality and neutralization studies both found that the main contributor to lethality without pretreatment with trypsin was CPB, whereas neutralization studies indicated that CPB and ETX were both important after trypsin pretreatment. At least part of the CPB produced by type B isolates remained active after trypsin treatment. However, the overall lethalities of most supernatants were lower after trypsin pretreatment. Also, there was a significant association between ETX, CPB2, and CPA production in vitro among type B isolates. However, our results suggest that both CPB and ETX are likely the most important contributors to the pathogenesis of C. perfringens type B infections in domestic animals.
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PMID:Both epsilon-toxin and beta-toxin are important for the lethal properties of Clostridium perfringens type B isolates in the mouse intravenous injection model. 1721 Jun 66

Clostridium perfringens type B causes enteritis and enterotoxemia in domestic animals. By definition, these bacteria must produce alpha toxin (CPA), beta toxin (CPB) and epsilon toxin (ETX) although most type B strains also produce perfringolysin O (PFO) and beta2 toxin (CPB2). A recently identified Agr-like quorum-sensing (QS) system in C. perfringens controls all toxin production by surveyed type A, C, and D strains, but whether this QS is involved in regulating toxin production by type B strains has not been explored. Therefore, the current study introduced agrB null mutations into type B strains CN1795 and CN1793. Both type B agrB null mutants exhibited reduced levels of CPB, PFO, and CPA in their culture supernatants, and this effect was reversible by complementation. The reduced presence of CPB in culture supernatant involved decreased cpb transcription. In contrast, the agrB null mutants of both type B strains retained wild-type production levels of ETX and CPB2. In a Caco-2 cell model of enteritis, culture supernatants of the type B agrB null mutants were less cytotoxic than supernatants of their wild-type parents. However, in an MDCK cell in vitro model for enterotoxemic effects, supernatants from the agrB null mutants or wild-type parents were equally cytotoxic after trypsin activation. Coupling these and previous results, it is now evident that strain-dependent variations exist in Agr-like QS system regulation of C. perfringens toxin production. The cell culture results further support a role for trypsin in determining which toxins contribute to disease involving type B strains.
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PMID:Role of the Agr-like quorum-sensing system in regulating toxin production by Clostridium perfringens type B strains CN1793 and CN1795. 2268 20


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