Activation and control of factor VII by activated factor X and thrombin. Isolation and characterization of a single chain form of factor VII.
Factor VII purified as previously described, was found to consist of two polypeptide chains joined by disulfide bridges. We now report the isolation and 200,000-fold purification of a single chain form of Factor VII. This was accomplished by protecting the molecule against proteolysis by including benzamidine during the entire purification. The purification was essentially as previously reported except that barium cirtate was substituted for barium sulfate as an absorbant for Factor VII as it resulted in a 4-fold increase in yield. Single chain Factor VII is rapidly hydrolyzed by Factor Xa in the presence of calcium ions and phospholipids, and by thrombin, to a two-chain form which possesses at least 85 times the Factor VII clotting activity of the single chain species. The two-chain form of the enzyme requires tissue factor in order to activate Factor X. From the observed rates of activation of Factor VII by Xa in the presence of calcium ions and phospholipids, it was calculated that at approximately physiological concentration, Factor VII activity would increase at an initial rate of 20-fold per min; this reaction is sufficiently rapid to constitute a feedback control mechanism. The action of thrombin is approximately 40-fold slower under these conditions. Diisopropylphosphorofluoridate inactivates the single chain and two-chain forms of Factor VII at approximately equal rates. After inhibition, the single chain species could be cleaved but not activated by proteolysis.
D000818:Animals; D002417:Cattle; D004789:Enzyme Activation; D005167:Factor VII / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D005170:Factor X / Q000378:metabolism*; D006801:Humans; D007531:Isoflurophate / Q000494:pharmacology; D011485:Protein Binding; D011487:Protein Conformation; D013917:Thrombin / Q000378:metabolism*
D007531:Isoflurophate; D005167:Factor VII; D005170:Factor X; D013917:Thrombin