IDENTIFICATION OF NOVEL PEPTIDE ANTAGONISTS FOR VON-WILLEBRAND-FACTORBINDING TO THE PLATELET GLYCOPROTEIN IB RECEPTOR FROM A PHAGE EPITOPELIBRARY

We have constructed a fusion phage epitope library in the filamentous bacteriophage fuse5. The library was made by inserting a degenerate ol igonucleotide which encodes 15 variable amino acids into the NH2-termi nal region of the phage geneIII protein. This library, containing over 10(7) different epitope bearing phage, has been used in an attempt to identify inhibitors of the von Willebrand factor (vWF)-platelet Glyco protein Ib interaction. The library was screened with a monoclonal ant ibody (RG46) that recognizes the GPIb binding domain of vWF (amino aci ds 445-733). A total of 30 clones falling into 8 classes have been ide ntified that react with the RG46 antibody. Isolates from all 8 classes are positive by immunoblot analysis. The amino acid sequence of the g ene III fusion protein from positive clones showed a strong homology t o the known RG46 epitope. Peptides identified from the screen were syn thesized and used to demonstrate that some of the synthetic peptides e xhibited inhibitory activity towards ristocetin induced binding of VWF to the GPIb receptor. Thus, we have demonstrated that screening a fus ion phage epitope library with a monoclonal antibody that inhibits vWF binding to the GPIb receptor can be a useful tool not only for mappin g antibody recognizing: determinants, but also can serve as a source f or identifying novel peptides that are antagonists for vWF binding to the platelet GPIb receptor.