Use of a neutralizing antibody helps identify structural features critical for binding of Clostridium difficile toxin TcdA to the host cell surface
| Autor: | D. Borden Lacy, Heather K. Kroh, Benjamin W. Spiller, Melanie D. Ohi, G. Jonah A. Rainey, Kim Rosenthal, Robert M. Woods, Paul Warrener, Ramyavardhanee Chandrasekaran, Xiaofang Jin, Andrew C. Nyborg |
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| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
Host cell surface biology medicine.drug_class 030106 microbiology Clostridium difficile toxin A Cell Biology Monoclonal antibody Endocytosis Biochemistry Virology Epitope Microbiology 03 medical and health sciences 030104 developmental biology Cell surface receptor medicine biology.protein Antibody Neutralizing antibody Molecular Biology |
| Zdroj: | Journal of Biological Chemistry. 292:14401-14412 |
| ISSN: | 0021-9258 |
| Popis: | Clostridium difficile is a clinically significant pathogen that causes mild-to-severe (and often recurrent) colon infections. Disease symptoms stem from the activities of two large, multidomain toxins known as TcdA and TcdB. The toxins can bind, enter, and perturb host cell function through a multistep mechanism of receptor binding, endocytosis, pore formation, autoproteolysis, and glucosyltransferase-mediated modification of host substrates. Monoclonal antibodies that neutralize toxin activity provide a survival benefit in preclinical animal models and prevent recurrent infections in human clinical trials. However, the molecular mechanisms involved in these neutralizing activities are unclear. To this end, we performed structural studies on a neutralizing monoclonal antibody, PA50, a humanized mAb with both potent and broad-spectrum neutralizing activity, in complex with TcdA. Electron microscopy imaging and multiangle light-scattering analysis revealed that PA50 binds multiple sites on the TcdA C-terminal combined repetitive oligopeptides (CROPs) domain. A crystal structure of two PA50 Fabs bound to a segment of the TcdA CROPs helped define a conserved epitope that is distinct from previously identified carbohydrate-binding sites. Binding of TcdA to the host cell surface was directly blocked by either PA50 mAb or Fab and suggested that receptor blockade is the mechanism by which PA50 neutralizes TcdA. These findings highlight the importance of the CROPs C terminus in cell-surface binding and a role for neutralizing antibodies in defining structural features critical to a pathogen's mechanism of action. We conclude that PA50 protects host cells by blocking the binding of TcdA to cell surfaces. |
| Databáze: | OpenAIRE |
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