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Figure 1. Identification of peptide-based inhibitors of Stx2 using tetravalent peptide libraries. A) The tetravalent peptide library was comprised of compounds with four randomized peptides of sequence Met-Ala-X-X-X-X-Ala-U (U; amino hexanoic acid), where X indicates all amino acids except Cys. B) The subset of tetravalent peptides that preferentially bound to 2BH- or 2BH-W33A-column was eluted and sequenced. The abundance of each amino acid in the peptides recovered from the 2BH-W33A-column, in a given cycle, was subtracted from the data obtained from the 2BH-column. Tetravalent peptide libraries with fixed Arg and/or Asn were used for the second round of selections. Values in parentheses indicate the relative selectivities for the amino acids. Boldface letters indicate amino acids that are strongly selected. C, D) Kinetics of the binding of each monomer peptide, the free trisaccharide analog (C), or the tetravalent peptide (D) to immobilized 2BH were analyzed using the BIAcoreTM system. PPR-mono, MAPPRRNRRA; PPP-mono, MAPPPRRRRA; KRR-mono, MAKRRNPRRA; FRR-mono, MAFRRNRRNA. The concentration of each tetravalent peptide is shown as the micromolar amount of the monomer peptide. (—) Binding was not detected.
