Activation of Purified Prothrombin in Ammonium Sulfate Solutions: Purification of Autoprothrombin C

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Abstract
For the generation of autoprothrombin C activity from prothrombin preparations in concentrated ammonium sulfate solution the optimum conditions were found to be near 2 M at pH 7. In addition to thrombin and autoprothrombin C an inhibitor was obtained. Methods were developed to obtain the autoprothrombin C consistently with a specific activity of about 4,700 units per mg protein, and a yield of one mg per liter of bovine plasma. At an intermediate stage of purification the stability of autoprothrombin C was far better than in the final product which lost activity even by simple freezing and thawing. Preservation of activity in 50% glycerol solutions at pH 7.2 was found to be the most convenient procedure. The alterations produced in purified prothrombin with DEAE cellulose chromatography are discussed with respect to possible significance for thrombin and autoprothrombin C. * This investigation was supported by a research grant HE 03424-08 from the National Heart Institute, National Institutes of Health, U.S. Public Health Service. Large quantities of bovine plasma were provided by Parke, Davis and Co., and we thank especially Mr. Thurston Springett of that company for keeping us supplied.