This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Nakao, H. Articles by Takeda, T. Search for Related Content PubMed PubMed Citation Articles by Nakao, H. Articles by Takeda, T.

 Previous Article  |  Next Article 

Infection and Immunity, November 1999, p. 5717-5722, Vol. 67, No. 11
0019-9567/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Monoclonal Antibody to Shiga Toxin 2 Which Blocks Receptor Binding and Neutralizes Cytotoxicity

Hiroshi Nakao,¹ Nobutaka Kiyokawa,² Junichiro Fujimoto,² Shinji Yamasaki,³ and Tae Takeda^1,*

Department of Infectious Diseases Research¹ and Department of Pathology and Pathophysiology,² National Children's Medical Research Center, Tokyo 154-8509, and Research Institute, International Medical Center of Japan, Tokyo 162-8655,³ Japan

Received 19 May 1999/Returned for modification 14 July 1999/Accepted 19 August 1999

A monoclonal antibody (MAb) was raised against Shiga toxin 2 (Stx2) of Escherichia coli O157:H7. MAb VTm1.1 belonged to the immunoglobulin G1 subclass and had a  light chain, and it could neutralize the cytotoxic activity of Stx2 and variants derived from patient strains but not that of variants derived from animals. MAb VTm1.1 was shown to bind to the B subunit of these neutralized Stx2s by Western blotting. Comparison of B-subunit amino acid sequences and reactivities to these Stxs suggested six amino acids (Ser30, Ser53, Glu56, Gln65, Asn68, and Asp69) that were candidates for the MAb VTm1.1 epitope. Consequently, five Stx2 mutants (S30N, S53N, E56H, Q65K, and N68Ter) were prepared by site-directed mutagenesis to determine which residue is essential for the epitope. All of these mutants showed cytotoxicity almost equal to that of the wild-type Stx2. Of the five Stx2 mutants, only E56H could not be neutralized by MAb VTm1.1. Western blot analysis also showed that MAb VTm1.1 could not bind to the E56H B subunit. These results indicated that Glu56 is an important residue recognized by MAb VTm1.1. Immunofluorescence analysis further indicated that MAb VTm1.1 inhibits the binding of Stx2 to its receptors. MAb VTm1.1 could be a useful therapeutic agent for Shiga toxin-producing E. coli infection.

---

^* Corresponding author. Mailing address: Department of Infectious Diseases Research, National Children's Medical Research Center, 3-35-31 Taishido, Setagaya-ku, Tokyo 154-8509, Japan. Phone: 81-3-3414-8121, ext. 2753. Fax: 81-3-3411-7308. E-mail: tae{at}nch.go.jp.

---

Infection and Immunity, November 1999, p. 5717-5722, Vol. 67, No. 11
0019-9567/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

This article has been cited by other articles:

• Smith, M. J., Carvalho, H. M., Melton-Celsa, A. R., O'Brien, A. D. (2006). The 13C4 Monoclonal Antibody That Neutralizes Shiga Toxin Type 1 (Stx1) Recognizes Three Regions on the Stx1 B Subunit and Prevents Stx1 from Binding to Its Eukaryotic Receptor Globotriaosylceramide. Infect. Immun. 74: 6992-6998 [Abstract] [Full Text]  
• Leung, P. H. M., Peiris, J. S. M., Ng, W. W. S., Yam, W. C. (2002). Polyclonal Antibodies to Glutathione S-Transferase- Verotoxin Subunit A Fusion Proteins Neutralize Verotoxins. CVI 9: 687-692 [Abstract] [Full Text]  
• Mukherjee, J., Chios, K., Fishwild, D., Hudson, D., O'Donnell, S., Rich, S. M., Donohue-Rolfe, A., Tzipori, S. (2002). Human Stx2-Specific Monoclonal Antibodies Prevent Systemic Complications of Escherichia coli O157:H7 Infection. Infect. Immun. 70: 612-619 [Abstract] [Full Text]