The Regulation of Clotting Factors

Blood clotting involves a multitude of proteins that act in concert in response to vascular injury to produce the procoagulant enzyme alpha-thrombin, which in turn is responsible for the generation of the fibrin plug. However, while generation of the fibrin plug is required for the arrest of excessive bleeding, unregulated clotting will result in the occlusion of the blood vessels and thrombosis. Thus, the regulation of the delicate balance between the procoagulant and anticoagulant mechanisms is of extreme importance for survival. While the majority of proteins involved in blood coagulation circulate as inactive zymogens that require proteolytic activation in order to function, approximately 1% of the circulating factor VII molecules are active. Factor VIIa, possess a serine protease active site, has poor catalytic activity, and is not inhibited by the circulating stoichiometric protease inhibitors. Following injury to the vasculature and subsequent exposure of the integral membrane glycoprotein, tissue factor (TF), the circulating factor VIIa molecules can bind to the exposed TF forming the extrinsic tenase complex (TF/factor VIIa) and initiate the blood coagulation process. Formation of the TF/factor VIIa complex increases the catalytic efficiency of the enzyme by four orders of magnitude when compared with factor VIIa alone. This cell-associated enzymatic complex initiates a series of enzymatic reactions, leading to the generation of alpha-thrombin and ultimately to the formation of the fibrin plug. The procoagulant enzymatic complexes (i.e., prothrombinase, intrinsic tenase, and extrinsic tenase) are similar in structure and composed of an enzyme, a cofactor, and the substrate associated on a cell surface in the presence of divalent metal ions. While the activity of the extrinsic tenase complex is limited by the availability (exposure) of its cell-associated cofactor (TF) it is remarkable that the activities of both the prothrombinase complex (factor Va/factor Xa) as well as the intrinsic tenase complex (factor VIIIa/factor IXa) are limited by the presence of the two soluble, nonenzymatic cofactors, factor Va and factor VIIIa. Factor Va and factor VIIIa, which are very similar in structure and function, are required for prothrombinase and intrinsic tenase activities, respectively, because both cofactors express a dual function in their respective complexes, acting as an enzyme receptor and catalytic effector on the cell surface. The cofactors derive from inactive plasma precursors by regulatory proteolytic events that involve alpha-thrombin. In general, bleeding tendencies are usually associated with defects in the activation of one of the zymogens or the cofactors of the procoagulant complexes. However, the activity of all of the complexes is also limited by the availability of an adequate membrane surface provided by endothelial cells, platelets, and monocytes. The cell surface provides a site for the recruitment of the appropriate proteins and allows for fast and efficient clot formation. In the absence of an appropriate membrane surface, the procoagulant complexes have limited catalytic efficiency. Thus, timely exposure of the adequate membrane surface is an additional step in the regulation of alpha-thrombin formation. alpha-Thrombin participates in its own down-regulation by binding to the endothelial cell receptor thrombomodulin, initiating the protein C pathway, which in turn leads to the formation of activated protein C (APC). APC is required for efficient neutralization of factor Va cofactor activity, which results in the inactivation of the prothrombin-activating complex. This inactivation can only occur in the presence of the appropriate membrane surface. Thus, while following alpha-thrombin activation, factor VIIIa is rapidly and spontaneously inactivated by dissociation of the A2 domain from the rest of the cofactor, APC is required for down-regulation of alpha-thrombin formation by prothrombinase. (ABSTRACT