Development peptide-based compounds which bind to the specific receptor-binding region of Shiga toxin B subunit

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To identify the peptide probes that specifically bind to one of the receptor-binding site 2 of Shiga toxin1 (Stx1), we established a novel screening system using the multivalent SPOT peptide array. We synthesized total 361 SPOT peptides on the cellulose membrane, and screened the peptides based on the binding affinities against the Stx1B subunit or Stx1 site 2 mutant (G62A). Finally, we identified 11 peptides that specifically bound to the Stx1B subunit, but not to G62A mutant. Furthermore, we found the 4 peptides among them (PQA-tet, KGA-tet, VIA-tet, YTA-tet) effectively inhibited the cytotoxicity for Stx1 in Vero cells. Thus, the multivalent SPOT peptide array technique may provide a powerful tool to identify a series of effective peptide-based Stx neutralizers which target a specific receptor binding region.