Abstract
Clostridioides difficile is a Gram-positive, pathogenic bacterium and a prominent cause of hospital-acquired diarrhea in the United States. The symptoms of C. difficile infection are caused by the activity of three large toxins known as toxin A (TcdA), toxin B (TcdB), and the C. difficile transferase toxin (CDT). Reported here is a 3.8-Å cryo-electron microscopy (cryo-EM) structure of CDT, a bipartite toxin comprised of the proteins CDTa and CDTb. We observe a single molecule of CDTa bound to a CDTb heptamer. The formation of the CDT complex relies on the interaction of an N-terminal adaptor and pseudoenzyme domain of CDTa with six subunits of the CDTb heptamer. CDTb is observed in a preinsertion state, a conformation observed in the transition of prepore to β-barrel pore, although we also observe a single bound CDTa in the prepore and β-barrel conformations of CDTb. The binding interaction appears to prime CDTa for translocation as the adaptor subdomain enters the lumen of the preinsertion state channel. These structural observations advance the understanding of how a single protein, CDTb, can mediate the delivery of a large enzyme, CDTa, into the cytosol of mammalian cells.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 6139-6144 |
| Number of pages | 6 |
| Journal | Proceedings of the National Academy of Sciences of the United States of America |
| Volume | 117 |
| Issue number | 11 |
| DOIs | |
| State | Published - Mar 17 2020 |
| Externally published | Yes |
Bibliographical note
Funding Information:The authors gratefully acknowledge James Fitzpatrick and Michael Rau (Washington University Center for Cellular Imaging) and Scott Collier and Melissa Chambers (Vanderbilt University Cryo-EM Facility) for their help in cryo-EM sample preparation and data collection. We thank members of the D.B.L. laboratory for their thoughtful feedback during manuscript preparation. This work was supported by U.S. Department of Veterans Affairs Award BX002943, Public Health Service Grant AI095755 from the National Institutes of Health, and Vanderbilt University. M.J.S. was supported by the Gastroenterology training program (DK007673). All molecular graphics and model analyses was performed with UCSF Chimera which was developed by the Resource for Biocomputing, Visualization, and Informatics at University of California, San Francisco, with support from NIH P41-GM103311.
Funding Information:
ACKNOWLEDGMENTS. The authors gratefully acknowledge James Fitzpatrick and Michael Rau (Washington University Center for Cellular Imaging) and Scott Collier and Melissa Chambers (Vanderbilt University Cryo-EM Facility) for their help in cryo-EM sample preparation and data collection. We thank members of the D.B.L. laboratory for their thoughtful feedback during manuscript preparation. This work was supported by U.S. Department of Veterans Affairs Award BX002943, Public Health Service Grant AI095755 from the National Institutes of Health, and Vanderbilt University. M.J.S. was supported by the Gastroenterology training program (DK007673). All molecular graphics and model analyses was performed with UCSF Chimera which was developed by the Resource for Biocomputing, Visualization, and Informatics at University of California, San Francisco, with support from NIH P41-GM103311.
Publisher Copyright:
© 2020 National Academy of Sciences. All rights reserved.
Keywords
- Binary toxin
- Clostridium
- Cryo-EM
- Iota toxin
