CHARACTERIZATION OF THE INITIAL ALPHA-THROMBIN INTERACTION WITH GLYCOPROTEIN IB-ALPHA IN RELATION TO PLATELET ACTIVATION

We have evaluated the properties of alpha-thrombin interaction with pl atelets within 1 min from exposure to the agonist, a time frame during which most induced activation responses are initiated and completed. Binding at 37 degrees C was rapidly reversible and completely blocked by a monoclonal antibody, LJ-Ib10, previously shown to be directed aga inst the alpha-thrombin interaction site on glycoprotein (GP) Ib alpha . By 2-5 min, however, binding was no longer fully reversible and was only partially inhibited by: the anti-GP Ib alpha antibody. Results we re similar at room temperature (22-25 degrees C), whereas the initial characteristics of alpha-thrombin interaction with platelets were pres erved for at least 20 min at 4 degrees C. Equilibrium binding isotherm s obtained at the latter temperature were compatible with a two-site m odel, but the component ascribed to GP IB alpha, 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 t he order of 10(-10) M) and low capacity as currently thought, The para meters of alpha-thrombin binding to intact GP Ib alpha on platelets at 4 degrees C corresponded closely to those measured with isolated GP I b alpha fragments regardless of temperature. Blocking the alpha-thromb in-GP Ib alpha interaction caused partial inhibition of ATP release an d prevented the association with platelets of measurable proteolytic a ctivity. These results support the concept that GP Ib alpha contribute s to the thrombogenic potential of alpha-thrombin.