INACTIVATION OF FACTOR Va BY PIASMIN

The inactivation of Factor Va by plasmin was studied in the presence and absence of phospholipid vesicles and calcium ions. The action of plasmin resulted in a rapid loss of the ability of Factor Va to serve as a cofactor to Factor Xa , as judged by clotting assays and . direct assays of prothrombin activation using the fluorophore, dansylarginine N-(3-ethyl-1,5-pentanediyl) amide (DAPA) . The rate of Factor Va inactivation catalyzed by plasmin was markedly enhanced by the addition of phospholipid vesicles (PCPS), suggesting that the action of plasmin on Factor Va may be a membrane bound phenomena. Both Factor Xa and prothrombin were capable of protecting Factor Va from inactivation by plasmin. SDS-PAGE was utilized to correlate plasmin catalyzed proteolysis of Factor Va with the concomitant loss of activity. Data obtained with Factor Va and the isolated chains of the cofactor indicated that the light chain (E) was cleaved by plasmin to yield products similar to those obtained with Factor Xa and Activated Protein C (APC). The heavy chain (D) was found to be degraded by plasmin to produce proteolytic fragments distinct from those produced by Factor Xa and APC. The action of plasmin on single chain Factor V was notable for an initial, transient increase in total Factor V activity, followed by subsequent loss of activity, indicating the transient formation of active intermediates. SDS-PAGE analysis revealed the degradation of Factor V by plasmin to final, inactive products via several transient, higher molecular weight intermediates. These findings may be of some significance in pathophysiologic states in which systemic fibrinolysis may occur, possibly contributing to the depletion of clotting factors. The identification of such ongoing processes may eventually be facilitated by the observation that the degradation of Factor Va by plasmin leads to end products which may be unique to this interaction.

(Supported by NIH Grants HL-35058 and HL 34575)