Coordinate binding of factor Va and factor Xa to the unstimulated platelet.
The interaction of Factor Xa and Factor Va at the platelet surface was investigated by direct, simultaneous binding measurements of both proteins to platelets and by immunochemical and kinetic techniques. Binding measurements of 125I-Factor Xa and 131I-Factor Va to unstimulated platelets indicate that the amount of Factor Xa bound is proportional to the amount of Factor Va bound. At saturating concentrations of Factor Xa, the ratio of platelet-bound Factor Xa to platelet-bound Factor Va was unity. At saturating levels of Factor Va (1.2 X 10(-8) M), 2300 molecules of Factor Xa are bound to the platelet, whereas at an equivalent concentration of unactivated Factor V, only 800 molecules of Factor Xa are bound. In the absence of exogenous Factor V or Va, thrombin-treated washed platelets bound only 200 Factor Xa molecules per platelet, suggesting that under these conditions, platelet Factor Va is the limiting component. Monovalent Fab fragments of burro antibovine Factor V inhibit, to the same degree, the rate of thrombin generation and the binding of Factor Va and Factor Xa to the platelet surface. Anti-Factor V Fab decreased the extent of Factor Va and Factor Xa binding equivalently. When the interaction of Factor Xa with platelets is modeled as Factor Xa binding to platelet-bound Factor Va, double reciprocal plots are linear, yielding a stoichiometry of 1.04 and a dissociation constant of 6 X 10(-10) M. Kinetic experiments indicate the presence of approximately 900 functional Factor Va platelet sites (Kd = 1.5 to 2.2 X 10(-10) M). This number of functional sites is equivalent to the number (837 +/- 48) of Factor Va high affinity binding sites (Kd = 4.0 X 10(-10) M). These sites most likely represent the Factor Xa binding sites involved in the function of the prothrombinase complex at the platelet surface.
D000818:Animals; D001665:Binding Sites; D001792:Blood Platelets / Q000378:metabolism*; D002417:Cattle; D004789:Enzyme Activation; D005165:Factor V / Q000378:metabolism*; D015943:Factor Va; D005170:Factor X / Q000378:metabolism*; D015951:Factor Xa; D007700:Kinetics; D011485:Protein Binding; D011516:Prothrombin / Q000378:metabolism
D015943:Factor Va; D005165:Factor V; D011516:Prothrombin; D005170:Factor X; D015951:Factor Xa