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Infection and Immunity, May 2007, p. 2381-2390, Vol. 75, No. 5
0019-9567/07/$08.00+0     doi:10.1128/IAI.01737-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Identification of a Prepore Large-Complex Stage in the Mechanism of Action of Clostridium perfringens Enterotoxin{triangledown}

James G. Smedley III,1,2 Francisco A. Uzal,3 and Bruce A. McClane1,2*

Department of Molecular Genetics and Biochemistry,1 Molecular Virology and Microbiology Graduate Program, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania,2 California Animal Health and Food Safety Laboratory, University of California, Davis, Davis, California3

Received 31 October 2006/ Returned for modification 26 December 2006/ Accepted 7 February 2007

Clostridium perfringens enterotoxin (CPE) is the etiological agent of the third most common food-borne illness in the United States. The enteropathogenic effects of CPE result from formation of large CPE-containing complexes in eukaryotic cell membranes. Formation of these ~155- and ~200-kDa complexes coincides with plasma membrane permeability changes in eukaryotic cells, causing a Ca2+ influx that drives cell death pathways. CPE contains a stretch of amino acids (residues 81 to 106) that alternates markedly in side chain polarity (a pattern shared by the transmembrane domains of the ß-barrel pore-forming toxin family). The goal of this study, therefore, was to investigate whether this CPE region is involved in pore formation. Complete deletion of the CPE region from 81 to 106 produced a CPE variant that was noncytotoxic for Caco-2 cells and was unable to form CPE pores. However, this variant maintained the ability to form the ~155-kDa large complex. This large complex appears to be a prepore present on the plasma membrane surface since it showed greater susceptibility to proteases, increased complex instability, and a higher degree of dissociation from membranes compared to the large complex formed by recombinant CPE. When a D48A mutation was engineered into this prepore-forming CPE variant, the resultant variant was unable to form any prepore ~155-kDa large complex. Collectively these findings reveal a new step in CPE action, whereby receptor binding is followed by formation of a prepore large complex, which then inserts into membranes to form a pore.

* Corresponding author. Mailing address: E1240 BSTWR, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261. Phone: (412) 648-9022. Fax: (412) 624-1401. E-mail: bamcc{at}pitt.edu

{triangledown} Published ahead of print on 16 February 2007.

Editor: D. L. Burns

Infection and Immunity, May 2007, p. 2381-2390, Vol. 75, No. 5
0019-9567/07/$08.00+0     doi:10.1128/IAI.01737-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Caserta, J. A., Hale, M. L., Popoff, M. R., Stiles, B. G., McClane, B. A. (2008). Evidence that Membrane Rafts Are Not Required for the Action of Clostridium perfringens Enterotoxin. Infect. Immun. 76: 5677-5685 [Abstract] [Full Text]  
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  • Smedley, J. G. III, Saputo, J., Parker, J. C., Fernandez-Miyakawa, M. E., Robertson, S. L., McClane, B. A., Uzal, F. A. (2008). Noncytotoxic Clostridium perfringens Enterotoxin (CPE) Variants Localize CPE Intestinal Binding and Demonstrate a Relationship between CPE-Induced Cytotoxicity and Enterotoxicity. Infect. Immun. 76: 3793-3800 [Abstract] [Full Text]  


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