Characterization of the inhibition of fibrin assembly by fibrinogen fragment D

Abstract

Fragment D (MW 100,000) prepared from a terminal plasmin digest of human fibrinogen was isolated and used to study its effect on fibrin formation. Increasing amounts of fragment D added to a solution of fibrinogen and thrombin decrease the rigidity of the resultant gel (10% of control at 2 mol of fragment D/mol fibrinogen). Half-maximal inhibition is achieved at 1 mol of fragment D/nol fibrinogen for non-cross-linked clots and at 1/2 mol of fragment D/mol fibrinogen for cross-linked clots. Clottability decreases concomitantly with the rigidity. Only small amounts of fragment D (< 10% for non-cross-linked gels) are incorporated into the gel. Light-scattering shows an increase in the final fiber thickness at fragment D concentrations up to 2 mol of fragment D/mol fibrinogen, from 60 molecule/cross-section for the control to 120 molecule/cross-section. Higher fragment D concentrations lead to a decrease in the final fiber thickness. The limit fiber thickness is 8 nm, with a length of 80 nm, which is equivalent to a firbin trimer. On the basis of results of synthetic-substrate and fibrinopeptide-release assays, it is clear that thrombin inactivation is not responsible for this effect. These data suggest that fragment D may inhibit fibrin formation by blocking the bimolecular polymerization of activated fibrin monomer molecules to form protofibrils, although additional effects on subsequent assembly steps may also be involved.