Summary
Purified fibrinogen preparations were clotted under crosslinking conditions and the evolution of α polymers was examined by Western blotting (SDS-PAGE). Monoclonal antibodies that bind to two localized regions within the COOH-terminal portion of the (A)α chains of fibrinogen (F-103, Aα #259-276; F-102, Aα #540-554) were used for immunodetection. Three crosslinked components (100K, 168K, 210K) that each exhibited coincident F-102 and F-103 immunoreactivities were evident as early as γ dimer formation under the in vitro conditions employed. Initial events in the a chain crosslinking process appeared to include interactions between relatively intact chains and a variety of degraded ones that shared the structure Aα #1-276, but differed in the extent to which regions between residues — #276 and — #539 were preserved. Degraded fibrinogen molecules whose Aα chains all terminated before Aα #540-554 were isolated from purified fibrinogen preparations by immunoaffinity chromatography on F-102 Sepharose. Immunoblotting data obtained for the crosslinking capacity of these degraded molecules indicated that early crosslinked components (95 K, 205 K) could form even in the absence of intact a chain partners. This crosslinking, moreover, could progress to the polymer stage. These findings demonstrate that some early crosslinking activity is localized exclusively within regions NH2-terminal to Aα — #540-554 and suggest that fibrinogen molecules with partially degraded Aα chains, which are likely to be circulating under many pathophysiologic conditions, can undergo fibrin stabilization through crosslinking despite a loss of at least 70 COOH-terminal Aα chain residues.
Keywords
Alpha chain crosslinking - Crosslinked fibrin - Crosslinking sites - Monoclonal antibodies
