This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Google Scholar
Right arrow Articles by Smith, M. J.
Right arrow Articles by O'Brien, A. D.
PubMed
Right arrow PubMed Citation
Right arrow Articles by Smith, M. J.
Right arrow Articles by O'Brien, A. D.

 Previous Article  |  Next Article 

Infection and Immunity, July 2009, p. 2730-2740, Vol. 77, No. 7
0019-9567/09/$08.00+0     doi:10.1128/IAI.00005-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Monoclonal Antibody 11E10, Which Neutralizes Shiga Toxin Type 2 (Stx2), Recognizes Three Regions on the Stx2 A Subunit, Blocks the Enzymatic Action of the Toxin In Vitro, and Alters the Overall Cellular Distribution of the Toxin{triangledown}

Michael J. Smith, Angela R. Melton-Celsa, James F. Sinclair, Humberto M. Carvalho, Cory M. Robinson, and Alison D. O'Brien*

Department of Microbiology and Immunology, Uniformed Services University of the Health Sciences, 4301 Jones Bridge Road, Bethesda, Maryland 20814-4799

Received 2 January 2009/ Returned for modification 6 March 2009/ Accepted 17 April 2009

Monoclonal antibody (MAb) 11E10 recognizes the Shiga toxin type 2 (Stx2) A1 subunit. The binding of 11E10 to Stx2 neutralizes both the cytotoxic and lethal activities of Stx2, but the MAb does not bind to or neutralize Stx1 despite the 61% identity and 75% similarity in the amino acids of the A1 fragments. In this study, we sought to identify the segment or segments on Stx2 that constitute the 11E10 epitope and to determine how recognition of that region by 11E10 leads to inactivation of the toxin. Toward those objectives, we generated a set of chimeric Stx1/Stx2 molecules and then evaluated the capacity of 11E10 to recognize those hybrid toxins by Western blot analyses and to neutralize them in Vero cell cytotoxicity assays. We also compared the amino acid sequences and crystal structures of Stx1 and Stx2 for stretches of dissimilarity that might predict a binding epitope on Stx2 for 11E10. Through these assessments, we concluded that the 11E10 epitope is comprised of three noncontiguous regions surrounding the Stx2 active site. To determine how 11E10 neutralizes Stx2, we examined the capacity of 11E10/Stx2 complexes to target ribosomes. We found that the binding of 11E10 to Stx2 prevented the toxin from inhibiting protein synthesis in an in vitro assay but also altered the overall cellular distribution of Stx2 in Vero cells. We propose that the binding of MAb 11E10 to Stx2 neutralizes the effects of the toxin by preventing the toxin from reaching and/or inactivating the ribosomes.

* Corresponding author. Mailing address: Department of Microbiology and Immunology, Uniformed Services University of the Health Sciences, Room B4052, 4301 Jones Bridge Road, Bethesda, MD 20814-4799. Phone: (301) 295-3400. Fax: (301) 295-3773. E-mail: aobrien{at}usuhs.mil

{triangledown} Published ahead of print on 11 May 2009.

Editor: V. J. DiRita

Infection and Immunity, July 2009, p. 2730-2740, Vol. 77, No. 7
0019-9567/09/$08.00+0     doi:10.1128/IAI.00005-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»