Reactions of thrombin-serpin complexes with thrombospondin.
Activated platelets release proteins that form stable complexes with thrombin (J. J. Miller, P. C. Browne, and T. C. Detwiler, Biochem. Biophys. Res. Commun. 151, 9-15, 1988). A working model for the reaction (P. C. Browne, J. J. Miller, and T. C. Detwiler, Arch. Biochem. Biophys. 265, 534-538, 1988) includes a dissociable complex of thrombin with released platelet protease nexin, leading to formation of a nondissociable thrombin-nexin complex that then becomes disulfide linked to thrombospondin. This disulfide-linked complex is converted back to the thrombin-nexin complex by reduction of disulfide bonds. Results that allow elaboration on this model are presented. After longer periods of incubation or after incubation with higher concentrations of thrombin, the amount of thrombin complexed with thrombospondin exceeded the amount of thrombin-nexin complex recovered after reduction of disulfide bonds. When the reaction mixture included inhibitors of formation of the thrombin-nexin complex, a slow formation of the thrombin-thrombospondin complex was observed. It was concluded that there is a nexin-independent as well as the faster nexin-dependent disulfide linkage of thrombin to thrombospondin. Addition of thrombin-antithrombin III complexes to the supernatant solution of activated platelets also led to complexes with thrombospondin, demonstrating that serpins other than platelet protease nexin facilitate incorporation of thrombin into complexes with thrombospondin. By heparin affinity chromatography, it was shown that thrombin-nexin complexes dissociably associate with thrombospondin prior to formation of disulfide-linked complexes. These observations are incorporated into a more detailed model of the reaction.
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
D001792:Blood Platelets / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008970:Molecular Weight; D015539:Platelet Activation; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism*; D011485:Protein Binding; D015843:Serpins / Q000378:metabolism*; D013917:Thrombin / Q000378:metabolism*; D019699:Thrombospondins
D010980:Platelet Membrane Glycoproteins; D015843:Serpins; D019699:Thrombospondins; D013917:Thrombin