Purification and characterization of thrombin from camel plasma: interaction with camel tick salivary gland thrombin inhibitor.

Abstract

Background: Thrombin is the most important enzyme in the hemostatic process by permitting rapid and localized coagulation in case of tissue damage. Camel thrombin is the natural and proper target enzyme for the previously purified camel tick salivary gland thrombin inhibitor.

Results: In this study, the camel thrombin was purified homogenously in a single affinity chromatographic step on the heparin-agarose affinity column with a specific activity of 3242 NIH units/mg proteins. On SDS-PAGE, the purified camel thrombin contained two forms, 37 kDa α-thrombin and 28 kDa β-thrombin, and the camel prothrombin was visualized as 72 kDa. The camel thrombin Km value was found out as 60 µM of N-(p-Tosyl)-Gly-Pro-Arg-p-nitroanilide acetate and displayed its optimum activity at pH 8.3. The PMSF was the most potent inhibitor of camel thrombin. Camel tick salivary gland thrombin inhibitor has two binding sites on camel thrombin and inhibited it competitively with Ki value of 0.45 µM.

Conclusions: The purified camel thrombin was found to be more susceptible toward the camel tick salivary gland thrombin inhibitor than bovine thrombin.