N-terminal Glycine Analysis for the Determination of the Proteolytic Activity of Thrombin^[*]

Summary

In the method presented in this paper for the assay of thrombin activity, N-terminal amino acids are determined with the phenylisothiocyanate method of Edman. Highly purified bovine fibrinogen is proteolyzed by thrombin under strictly defined conditions. The increase in N-terminal glycine is then directly proportional to the activity of the thrombin. Five different thrombin preparations were standardized with this N-terminal method and with a clotting method. The results obtained with the two methods are in good agreement. Because the N-terminal method is accurate and highly specific for thrombin it is proposed as standard method for the chemical determination of thrombin activity. One NIH unit of thrombin was found to make 0.4 μimoles of glycine appear in N-terminal position.

^* Supported by grants from the Swedish National Association against Tuberculosis and other Social Diseases and from the Swedish Society for Medical Research.

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