PURIFICATION AND SOME PROPERTIES OF AUTOPROTHROMBIN C

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Abstract
Methods were developed for the isolation of autoprothrombin C, which is the second enzyme obtained from purified bovine prothrombin. In the presence of lipids and standardized conditions 0.35 μg of the purified autoprothrombin C were sufficient for clotting recalcified plasma in 15 seconds. Some physicochemical properties are as follows: S20, w is 2.27S, the diffusion constant 8.4 × 10−7 cm2/second, and the partial specific volume 0.695. The molecular weight from physicochemical measurements was 21,500. All amino acids were found, but only 1 molecule of methionine. On the basis of amino acid composition the molecular weight was found to be 27,000, giving an average of 24,200 for our two determinations. Prothrombin contains sufficient of each amino acid residue to supply autoprothrombin C and thrombin. Autoprothrombin II, however, has only sufficient amino acids for either autoprothrombin C or thrombin, but not both. The purified autoprothrombin C contained 7% carbohydrate (orcinol) and 3.8% hexosamine. It was stable at pH 7.2 for more than a week at room temperature, and longer in subzero glycerol solution. Autoprothrombin C was used to obtain a single precipitin band in agar diffusion plates with antibody to prothrombin. The band also identified with a single plasma antibody.