CHARACTERISTICS OF THE INTERACTION BETWEEN THROMBIN EXOSITE-1 AND THESEQUENCE-269-297 OF PLATELET GLYCOPROTEIN IB-ALPHA

The interaction between GPIb and thrombin promotes platelet activation elicited via the hydrolysis of the thrombin receptor and involves str uctures located on the segment 238-290 within the N-terminal domain of GPIb alpha and the positively charged exosite 1 on thrombin. We have investigated the ability of peptides derived from the 269-287 sequence of GPIb alpha to interact with thrombin. Three peptides were synthesi zed, including Ib alpha 269-287 and two scrambled peptides R1 and R2 w hich are comparable to Ib alpha 269-287 with regards to their content and distribution of anionic residues. However, R2 differs from both Ib alpha 269-287 and R1 by the shifting of one proline from a central po sition to the N-terminus. By chemical cross-linking, we observed the f ormation of a complex between I-125-Ib alpha 269-287 and alpha-thrombi n that was inhibited by hirudin, the C-terminal peptide of hirudin, so dium pyrophosphate but not by heparin. The complex did not form when g amma-thrombin was substituted for alpha-thrombin. Ib alpha 269-287 pro duced only slight changes in thrombin amidolytic activity and inhibite d thrombin binding to fibrin. R1 and R2 also formed complexes with alp ha-thrombin, modified slightly its catalytic activity and inhibited it s binding to fibrin. Peptides Ib alpha 269-287 and R1 inhibited platel et aggregation and secretion induced by low thrombin concentrations wh ereas R2 was without effect. Our results indicate that Ib alpha 269-28 7 interacts with thrombin exosite 1 via mainly electrostatic interacti ons, which explains why the scrambled peptides also interact with exos ite 1. Nevertheless, the lack of effect of R2 on thrombin-induced plat elet activation suggests that proline 280 is important for thrombin in teraction with GPIb.