Isolation and Purification of a Coagulant from Snake Venom of the Species Bothrops jararaca and the Study of its Properties

Summary

1. A material active in coagulating fibrinogen has been purified thirty-fold from crude venom of the species Bothrops jararaca. This material exhibits clotting and esterase activities, but no measurable proteolytic activity if measured by its ability to digest denatured hemoglobin.

2. The clotting activity of the purified material (but not its esterase activity) is greatly enhanced by the addition of 2—3 μM of calcium ion; no other divalent metallic ion can duplicate the accelerating effect of Ca⁺⁺ ion.

3. The purified material is stable to dialysis, standing at room temperature for 18—20 hours, or heating at 50° C for 10 minutes; it loses both its clotting and esterase activities if heated at 70° C for 25 minutes or at 80° C for 10 minutes

4. Although the powerful proteolytic enzyme inhibitors ovomucoid and soyabean trypsin inhibitor can block the clotting and esterase activities of thrombin, they are ineffective with respect to these properties of venom-coagulin. DFP, an inhibitor of far less specificity, can prevent the actions both of thrombin and of venom-coagulin

5. An amount of heparin sufficient to block the clotting activity of thrombin is unable to prevent the action of venom-coagulin on fibrinogen; a much higher concentration of heparin is needed. In both cases the addition of 2—3 μM of Ca⁺⁺ ion necessitates the introduction of considerably more heparin in order to block the actions of thrombin and venom-coagulin.

6. While the antithrombin contained in serum can neutralize the clotting and esterase activities of thrombin, it has no effect on the actions of venom-coagulin.

^* Present addresses: 1. Department of Biochemistry, North-Western University Medical School, Chicago, Illinois; 2. Department of Biochemistry, College of Physicians and Surgeons, New York, New York; 3. Department of Biochemistry, Rochester University Medical School, Rochester, New York.

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