The reaction of thrombin with platelet-derived nexin requires a secondary recognition site in addition to the catalytic site.
A protease nexin released by activated platelets forms stable complexes with alpha-thrombin. Active-site-blocked thrombin does not form the stable complex, but it inhibits formation of the stable complex by active alpha-thrombin. gamma-Thrombin, which has a damaged substrate recognition site (the anion-binding exosite), did not form the complex and did not inhibit formation of the stable complex by alpha-thrombin. Complex formation was inhibited by the C-terminal dodecapeptide of hirudin, which has been shown to bind to the anion-binding exosite. A monoclonal antibody that blocks reactions of thrombin that involve the anion-binding exosite also inhibited formation of a stable complex of alpha-thrombin and the platelet-derived protease nexin. It is concluded that the anion-binding exosite of thrombin, a site that confers a high degree of specificity for substrates with a complementary site, binds to the platelet nexin prior to reaction of the catalytic site with the serpin.
Chang
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AC
Department of Biochemistry, State University of New York Health Science Center, Brooklyn 11203.
Detwiler
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TC
D016564:Amyloid beta-Protein Precursor; D000911:Antibodies, Monoclonal; D001665:Binding Sites; D001792:Blood Platelets / Q000378:metabolism*; D000001:Calcimycin / Q000494:pharmacology; D002352:Carrier Proteins / Q000097:blood* / Q000302:isolation & purification; D006801:Humans; D007700:Kinetics; D015539:Platelet Activation; D058255:Protease Nexins; D011485:Protein Binding; D011956:Receptors, Cell Surface; D013917:Thrombin / Q000037:antagonists & inhibitors / Q000378:metabolism*
D016564:Amyloid beta-Protein Precursor; D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; D058255:Protease Nexins; D011956:Receptors, Cell Surface; D000001:Calcimycin; D013917:Thrombin