Purification and characterisation of Clostridium difficile toxin A by bovine thyroglobulin affinity chromatography and dissociation in denaturing conditions with or without reduction - PubMed (original) (raw)
Purification and characterisation of Clostridium difficile toxin A by bovine thyroglobulin affinity chromatography and dissociation in denaturing conditions with or without reduction
S Kamiya et al. J Med Microbiol. 1989 Sep.
Abstract
Highly purified toxin A of Clostridium difficile was obtained by bovine thyroglobulin affinity chromatography followed by two sequential anion-exchange chromatography steps on Q Sepharose FF and Mono Q. After Q Sepharose FF chromatography of a thyroglobulin affinity-purified toxin A preparation, two major peaks of cytotoxicity representing toxins A and B were detected. The homogeneity of the final toxin A preparation obtained from Mono Q anion-exchange fast protein liquid chromatography was ascertained by gel electrophoresis developed by silver stain. The mol. wt of toxin A in non-denaturing conditions was estimated to be 520-540 Kda by native polyacrylamide gel electrophoresis (PAGE) developed by silver stain. In contrast, with sodium dodecyl sulphate (SDS)-PAGE under reducing or non-reducing conditions, a major band of 240 Kda and 10 minor and 27 faint bands (non-reduced conditions), or four minor and 31 faint bands (reduced conditions) were detected after silver staining. In two-dimensional PAGE, the seven minor bands of greater than 240 Kda obtained by non-reducing SDS-PAGE migrated to the 240-Kda position after reduction with beta-mercaptoethanol.
Similar articles
- A non-haemagglutinating form of Clostridium difficile toxin A.
Kamiya S, Borriello SP. Kamiya S, et al. J Med Microbiol. 1992 Mar;36(3):190-7. doi: 10.1099/00222615-36-3-190. J Med Microbiol. 1992. PMID: 1548692 - Purification and characterisation of intracellular toxin A of Clostridium difficile.
Meng XQ, Kamiya S, Yamakawa K, Ogura H, Nakamura S. Meng XQ, et al. J Med Microbiol. 1993 Jan;38(1):69-73. doi: 10.1099/00222615-38-1-69. J Med Microbiol. 1993. PMID: 8418298 - Purification of Clostridium difficile toxin A by affinity chromatography on immobilized thyroglobulin.
Krivan HC, Wilkins TD. Krivan HC, et al. Infect Immun. 1987 Aug;55(8):1873-7. doi: 10.1128/iai.55.8.1873-1877.1987. Infect Immun. 1987. PMID: 3112015 Free PMC article. - Simplified purification method for Clostridium difficile toxin A.
Fu SW, Xue J, Zhang YL, Zhou DY. Fu SW, et al. World J Gastroenterol. 2004 Sep 15;10(18):2756-8. doi: 10.3748/wjg.v10.i18.2756. World J Gastroenterol. 2004. PMID: 15309736 Free PMC article. - A review on native and denaturing purification methods for non-coding RNA (ncRNA).
Kanwal F, Lu C. Kanwal F, et al. J Chromatogr B Analyt Technol Biomed Life Sci. 2019 Jul 1;1120:71-79. doi: 10.1016/j.jchromb.2019.04.034. Epub 2019 Apr 26. J Chromatogr B Analyt Technol Biomed Life Sci. 2019. PMID: 31071581 Review.
Cited by
- Effects of Clostridium difficile toxin A on K562/A02 cell proliferation, apoptosis and multi-drug resistance.
Xi Y, Ma Z, Zhang H, Yuan M, Wang L. Xi Y, et al. Oncol Lett. 2018 Apr;15(4):4215-4220. doi: 10.3892/ol.2018.7921. Epub 2018 Feb 1. Oncol Lett. 2018. PMID: 29545897 Free PMC article. - Circulating antibody and memory B-Cell responses to C. difficile toxins A and B in patients with C. difficile-associated diarrhoea, inflammatory bowel disease and cystic fibrosis.
Monaghan TM, Robins A, Knox A, Sewell HF, Mahida YR. Monaghan TM, et al. PLoS One. 2013 Sep 10;8(9):e74452. doi: 10.1371/journal.pone.0074452. eCollection 2013. PLoS One. 2013. PMID: 24058568 Free PMC article. - Primary human colonic myofibroblasts are resistant to Clostridium difficile toxin A-induced, but not toxin B-induced, cell death.
Mullan N, Hughes KR, Mahida YR. Mullan N, et al. Infect Immun. 2011 Apr;79(4):1623-30. doi: 10.1128/IAI.00686-10. Epub 2011 Jan 18. Infect Immun. 2011. PMID: 21245273 Free PMC article. - Antibody-enhanced, Fc gamma receptor-mediated endocytosis of Clostridium difficile toxin A.
He X, Sun X, Wang J, Wang X, Zhang Q, Tzipori S, Feng H. He X, et al. Infect Immun. 2009 Jun;77(6):2294-303. doi: 10.1128/IAI.01577-08. Epub 2009 Mar 23. Infect Immun. 2009. PMID: 19307220 Free PMC article. - Essential role of toxin A in C. difficile 027 and reference strain supernatant-mediated disruption of Caco-2 intestinal epithelial barrier function.
Sutton PA, Li S, Webb J, Solomon K, Brazier J, Mahida YR. Sutton PA, et al. Clin Exp Immunol. 2008 Sep;153(3):439-47. doi: 10.1111/j.1365-2249.2008.03690.x. Epub 2008 May 26. Clin Exp Immunol. 2008. PMID: 18505424 Free PMC article.