The factor Xa-catalyzed activation of factor V.
Factor V appears to be a procofactor with, at best, 1/400 the activity of fully activated Factor V (Factor Va). The proteolytic conversion of Factor V to Factor Va is catalyzed by thrombin. However, since Factor Va activity is required for thrombin generation, the initial participation of Factor V in the expression of prothrombinase activity is not well understood. In the present study, the activation of Factor V by Factor Xa has been investigated. Cofactor activation was assessed by monitoring the conversion of prethrombin-1 to thrombin in the presence of 5-dimethylamino-naphthalene-1-sulfonylarginine-N-(3-ethyl-1,5-pentanediyl)amide (DAPA). The DAPA not only provided a fluorescent signal for the formation of thrombin, but also attenuated the feedback activation of Factor V by thrombin. Trace quantities of Factor Va were removed from the Factor V preparations by immunoadsorption with immobilized murine monoclonal antibodies selective for Factor Va. The incubation of Factor V with Factor Xa in the presence of phosphatidylcholine/phosphatidylserine vesicles, CaCl2, and DAPA resulted in a time-dependent increase in cofactor activity. Phosphatidylcholine/phosphatidylserine vesicles were not absolutely required, but the rate of Factor V activation was significantly enhanced by inclusion of the vesicles. The activation was absolutely dependent upon Factor Xa and was eliminated by immunoadsorption of the Factor Xa preparation with a murine anti-Factor X (Xa) monoclonal antibody coupled to agarose. The activation was not affected by immunoadsorption of the Factor Xa and Factor V preparations with burro polyclonal anti-prothrombin IgG. Most of the products of the Factor Xa activation of Factor V differ from the products derived by the thrombin-catalyzed activation of the procofactor. The results demonstrate that Factor Xa catalyzes the activation of Factor V. Furthermore, these studies suggest that the Factor Xa activation of Factor V may be responsible for the advent of early prothrombinase activity.
D000818:Animals; D002417:Cattle; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D005165:Factor V / Q000302:isolation & purification / Q000378:metabolism*; D005170:Factor X / Q000378:metabolism*; D015951:Factor Xa; D007700:Kinetics; D008081:Liposomes; D008970:Molecular Weight; D011516:Prothrombin / Q000378:metabolism
D008081:Liposomes; D005165:Factor V; D011516:Prothrombin; D005170:Factor X; D015951:Factor Xa