We have evaluated the properties of α-thrombin interaction with platelets within 1 min from exposure to the agonist, a time frame during which most induced activation responses are initiated and completed. Binding at 37 °C was rapidly reversible and completely blocked by a monoclonal antibody, LJ-Ib10, previously shown to be directed against the α-thrombin interaction site on glycoprotein (GP) Ibα. By 2-5 min, however, binding was no longer fully reversible and was only partially inhibited by the anti-GP Ibα antibody. Results were similar at room temperature (22-25 °C), whereas the initial characteristics of α-thrombin interaction with platelets were preserved for at least 20 min at 4 °C. Equilibrium binding isotherms obtained at the latter temperature were compatible with a two-site model, but the component ascribed to GP Ibα, completely inhibited by LJ-Ib10, had 'moderate' affinity (k(d) on the order of 10-8 M) and relatively high capacity, rather than 'high' affinity (k(d) on the order of 10-10 M) and low capacity as currently thought. The parameters of α-thrombin binding to intact GP Ibα on platelets at 4 °C corresponded closely to those measured with isolated GP Ibα fragments regardless of temperature. Blocking the α- thrombin-GP Ibα interaction caused partial inhibition of ATP release and prevented the association with platelets of measurable proteolytic activity. These results support the concept that GP Ibα contributes to the thrombogenic potential of α-thrombin.
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