Structural Basis for the Platelet-Collagen Interaction: THE SMALLEST MOTIF WITHIN COLLAGEN THAT RECOGNIZES AND ACTIVATES PLATELET GLYCOPROTEIN VI CONTAINS TWO GLYCINE-PROLINE-HYDROXYPROLINE TRIPLETS

Collagen-related peptide is a selective agonist for the platelet collagen receptor Glycoprotein VI. The triple helical peptide contains ten GPO triplets/strand (single letter amino acid nomenclature, where O is hydroxyproline) and so over-represents GPO compared with native collagen sequence. To investigate the ability of Glycoprotein VI to recognize GPO triplets in a setting more representative of the collagens, we synthesized a set of triple helical peptides containing fewer GPO triplets, varying their number and spacing within an inert (GPP)n backbone. The adhesion of recombinant human Glycoprotein VI ectodo-main, like that of human platelets, to these peptides increased with their GPO content, and platelet adhesion was abolished by the specific anti-Glycoprotein VI-blocking antibody, 10B12. Platelet aggregation and protein tyrosine phosphorylation were induced only by cross-linked peptides and only those that contained two or more GPO triplets. Such peptides were less potent than cross-linked collagen-related peptide. Our data suggest that both the sequences GPOGPO and GPO...GPO represent functional Glycoprotein VI recognition motifs within collagen. Furthermore, we propose that the (GPO)₄ motif can support simultaneous binding of two glycoprotein VI molecules, in either a parallel or anti-parallel stacking arrangement, which could play an important role in activation of signaling.