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Published January 2010 | public
Journal Article

Three-dimensional structure of the detergent-solubilized Vibrio cholerae cytolysin (VCC) heptamer by electron cryomicroscopy

Abstract

Vibrio cholerae cytolysin (VCC) is a pore-forming toxin that inserts a lytic water-filled channel into susceptible host membranes. Assembly of the toxin on cell surfaces may be enhanced by two tandem lectin domains, in addition to direct interactions with lipids and cholesterol within the membrane itself. We used single-particle electron cryomicroscopy (cryoEM) to generate a low-resolution molecular structure of the detergent-solubilized VCC oligomer to 20 Å resolution. After confirming a heptameric arrangement of individual protomers, sevenfold averaging around the central pore was utilized to improve the structure. Docking of the previously determined VCC protoxin crystal structure was possible with rigid-body rearrangements between the cytolytic and lectin domains. A second cryoEM reconstruction of a truncated VCC mutant supported the topology of our model in which the carboxyl-terminal lectin domain forms "spikes" around the toxin core with the putative carbohydrate receptor-binding site accessible on the surface of the oligomer. This finding points to an assembly mechanism in which lectin domains may remain bound to receptors on the cell surface throughout assembly of the cytolytic toxin core and explains the hemagglutinating activity of purified toxin. Our model provides an insight into the structural rearrangements that accompany VCC-mediated cytolysis and may aid in the engineering of novel pore-forming toxins to attack specific cells towards therapeutic ends.

Additional Information

© 2009 Elsevier Inc. Received 20 May 2009; revised 13 July 2009; accepted 14 July 2009. Available online 16 July 2009. We thank Pamela Björkman for her advice and support throughout this project. We also thank William Tivol, Alasdair McDowall, and Jian Shi for their help with the electron microscopy and Grant Jensen for reading our manuscript. Xavier Siebert provided assistance with the UROX software. This work was supported by a Beckman Institute Fellowship (R.O.), a postdoctoral fellowship from the Cancer Research Institute (Y.H.), and gifts to Caltech to support electron microscopy from the Gordon and Betty Moore Foundation and the Agouron Institute. Molecular graphics images were produced using the UCSF Chimera package from the Resource for Biocomputing, Visualization, and Informatics at the University of California, San Francisco (supported by NIH P41 RR-01081).

Additional details

Created:
August 21, 2023
Modified:
October 19, 2023