Isolation and characterization of thrombomodulin from bovine lung.
Bovine thrombomodulin was isolated from the lung by Triton X extraction, affinity chromatography on diisopropyl phosphate-thrombin-agarose, and gel filtration on Ultrogel AcA-44. The final preparation was purified 6000-fold from the membrane extract with a yield of 21%. It showed apparent Mr of 78,000 and 105,000, before and after reduction, respectively, on polyacrylamide gel electrophoresis in SDS. The activity of the thrombomodulin was stable under the conditions of 1% SDS, 8 M urea, pH 2 and 10, and heat treatment at 60 degrees C for 30 min, but was unstable against treatment with 2-mercaptoethanol. Activation of protein C by thrombin in the presence of the thrombomodulin depended on Ca2+, and an equimolar complex formation between thrombin and thrombomodulin was required for the maximum rate activation. The rate of protein C activation by thrombin was increased 900-fold by thrombomodulin. Thrombomodulin inhibited the thrombin-induced fibrinogen clotting and platelet activation. However, it did not affect the inhibition of thrombin by antithrombin III with or without heparin, a protein C inhibitor or several synthetic inhibitors. These properties of bovine thrombomodulin were similar to those of rabbit thrombomodulin reported earlier.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D000990:Antithrombin III / Q000494:pharmacology; D002417:Cattle; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D004789:Enzyme Activation; D006023:Glycoproteins / Q000378:metabolism; D006493:Heparin / Q000494:pharmacology; D006801:Humans; D008168:Lung / Q000032:analysis*; D008970:Molecular Weight; D010974:Platelet Aggregation / Q000187:drug effects; D011486:Protein C; D011817:Rabbits; D011956:Receptors, Cell Surface / Q000302:isolation & purification*; D018179:Receptors, Thrombin; D013045:Species Specificity; D013917:Thrombin / Q000378:metabolism
D000596:Amino Acids; D006023:Glycoproteins; D011486:Protein C; D011956:Receptors, Cell Surface; D018179:Receptors, Thrombin; D000990:Antithrombin III; D006493:Heparin; D013917:Thrombin