Identification of an Essential Region for Translocation of Clostridium difficile Toxin B
Clostridium difficile toxin A (TcdA) and toxin B (TcdB) are the major virulence factors involved in C. difficile-associated diarrhea and pseudomembranous colitis. TcdA and TcdB both contain at least four distinct domains: the glucosyltransferase domain, cysteine protease domain, receptor binding dom...
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| Format: | Article |
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MDPI AG
2016-08-01
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| Series: | Toxins |
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| Online Access: | http://www.mdpi.com/2072-6651/8/8/241 |
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| author | Shuyi Chen Haiying Wang Huawei Gu Chunli Sun Shan Li Hanping Feng Jufang Wang |
| author_facet | Shuyi Chen Haiying Wang Huawei Gu Chunli Sun Shan Li Hanping Feng Jufang Wang |
| author_sort | Shuyi Chen |
| collection | DOAJ |
| description | Clostridium difficile toxin A (TcdA) and toxin B (TcdB) are the major virulence factors involved in C. difficile-associated diarrhea and pseudomembranous colitis. TcdA and TcdB both contain at least four distinct domains: the glucosyltransferase domain, cysteine protease domain, receptor binding domain, and translocation domain. Few studies have investigated the translocation domain and its mechanism of action. Recently, it was demonstrated that a segment of 97 amino acids (AA 1756–1852, designated D97) within the translocation domain of TcdB is essential for the in vitro and in vivo toxicity of TcdB. However, the mechanism by which D97 regulates the action of TcdB in host cells and the important amino acids within this region are unknown. In this study, we discovered that a smaller fragment, amino acids 1756–1780, located in the N-terminus of the D97 fragment, is essential for translocation of the effector glucosyltransferase domain into the host cytosol. A sequence of 25AA within D97 is predicted to form an alpha helical structure and is the critical part of D97. The deletion mutant TcdB∆1756–1780 showed similar glucosyltransferase and cysteine protease activity, cellular binding, and pore formation to wild type TcdB, but it failed to induce the glucosylation of Rho GTPase Rac1 of host cells. Moreover, we found that TcdB∆1756–1780 was rapidly degraded in the endosome of target cells, and therefore its intact glucosyltransferase domain was unable to translocate efficiently into host cytosol. Our finding provides an insight into the molecular mechanisms of action of TcdB in the intoxication of host cells. |
| first_indexed | 2024-04-14T02:15:43Z |
| format | Article |
| id | doaj.art-b2f42a88a2ad4e129e8de1ea72c87cde |
| institution | Directory Open Access Journal |
| issn | 2072-6651 |
| language | English |
| last_indexed | 2024-04-14T02:15:43Z |
| publishDate | 2016-08-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Toxins |
| spelling | doaj.art-b2f42a88a2ad4e129e8de1ea72c87cde2022-12-22T02:18:13ZengMDPI AGToxins2072-66512016-08-018824110.3390/toxins8080241toxins8080241Identification of an Essential Region for Translocation of Clostridium difficile Toxin BShuyi Chen0Haiying Wang1Huawei Gu2Chunli Sun3Shan Li4Hanping Feng5Jufang Wang6School of Bioscience and Bioengineering, South China University of Technology, Guangzhou 510006, ChinaSchool of Bioscience and Bioengineering, South China University of Technology, Guangzhou 510006, ChinaSchool of Bioscience and Bioengineering, South China University of Technology, Guangzhou 510006, ChinaSchool of Bioscience and Bioengineering, South China University of Technology, Guangzhou 510006, ChinaSchool of Bioscience and Bioengineering, South China University of Technology, Guangzhou 510006, ChinaDepartment of Microbial Pathogenesis, University of Maryland Dental School, Baltimore, MD 21201, USASchool of Bioscience and Bioengineering, South China University of Technology, Guangzhou 510006, ChinaClostridium difficile toxin A (TcdA) and toxin B (TcdB) are the major virulence factors involved in C. difficile-associated diarrhea and pseudomembranous colitis. TcdA and TcdB both contain at least four distinct domains: the glucosyltransferase domain, cysteine protease domain, receptor binding domain, and translocation domain. Few studies have investigated the translocation domain and its mechanism of action. Recently, it was demonstrated that a segment of 97 amino acids (AA 1756–1852, designated D97) within the translocation domain of TcdB is essential for the in vitro and in vivo toxicity of TcdB. However, the mechanism by which D97 regulates the action of TcdB in host cells and the important amino acids within this region are unknown. In this study, we discovered that a smaller fragment, amino acids 1756–1780, located in the N-terminus of the D97 fragment, is essential for translocation of the effector glucosyltransferase domain into the host cytosol. A sequence of 25AA within D97 is predicted to form an alpha helical structure and is the critical part of D97. The deletion mutant TcdB∆1756–1780 showed similar glucosyltransferase and cysteine protease activity, cellular binding, and pore formation to wild type TcdB, but it failed to induce the glucosylation of Rho GTPase Rac1 of host cells. Moreover, we found that TcdB∆1756–1780 was rapidly degraded in the endosome of target cells, and therefore its intact glucosyltransferase domain was unable to translocate efficiently into host cytosol. Our finding provides an insight into the molecular mechanisms of action of TcdB in the intoxication of host cells.http://www.mdpi.com/2072-6651/8/8/241Clostridium difficiletoxintranslocationendocytosisconformational change |
| spellingShingle | Shuyi Chen Haiying Wang Huawei Gu Chunli Sun Shan Li Hanping Feng Jufang Wang Identification of an Essential Region for Translocation of Clostridium difficile Toxin B Toxins Clostridium difficile toxin translocation endocytosis conformational change |
| title | Identification of an Essential Region for Translocation of Clostridium difficile Toxin B |
| title_full | Identification of an Essential Region for Translocation of Clostridium difficile Toxin B |
| title_fullStr | Identification of an Essential Region for Translocation of Clostridium difficile Toxin B |
| title_full_unstemmed | Identification of an Essential Region for Translocation of Clostridium difficile Toxin B |
| title_short | Identification of an Essential Region for Translocation of Clostridium difficile Toxin B |
| title_sort | identification of an essential region for translocation of clostridium difficile toxin b |
| topic | Clostridium difficile toxin translocation endocytosis conformational change |
| url | http://www.mdpi.com/2072-6651/8/8/241 |
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