Experimental Studies on Fibrin Polymerization and Intrinsic Fibrinolysis

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It is essential for blood clot generation that fibrinogen is converted into fibrin as a result of thrombin cleaving peptides from the amino-terminal ends of the alpha and beta chains. However, the details required for a full mechanistic explanation of the fibrin formation including the polymerization sites has not been clarified until now.In this project, we investigated the characterization of the fibrin polymerization site in the amino-terminus of the beta chain utilizing synthetic peptides. The binding of the synthetic peptide, Gly-His-Arg-Pro-amide, which corresponds to the amino-terminus of the beta chain, to fragment D1 was found to depend on the concentration of calcium ions, being maximal at 10^<-4>M and decreasing on either side of this at room temperature. In contrast, the effect of calcium ions on the binding was different when examined using a fragment D1 sepharose column treated with sialidase. These results emphasize the close proximity of the calcium-binding and peptide-binding sites in the calboxy-terminus of beta chain to which the carbohydrate attaches. Furthermore, we found the synthetic peptide Gly-His-Arg to have a slight protective effect on the gamma chain remnant of fragment D1 against plasmin in the same fashion as calcium ions.These date indicate that one of the binding sites of the amino-terminal ends of beta chain is located in the carboxy-terminus of beta chain and the other is located in the carboxy-terminus of gamma chain.