Measurements of clot rigidity and fiber thickness indicate that fragment D is a potent inhibitor of fibrin assembly. At physiological ionic strength, D concentrations in excess of 2 moles D/mole fibrinogen lead to a large decrease in clot rigidity and fiber thickness. Above 14 moles D/mole fibrinogen, gelation is inhibited. The molecular weight and radius of gyration of the fibers, determined by light scattering, confirm that short oligomers result, composed of 3 fibrin monomer molecules at 120 moles D/mole fibrinogen and 9 monomers at 14.4 moles D/mole fibrinogen. If D is added to a solution of long protofibrils, no inhibition is observed. Apparently the polymerization of monomers to protofibrils is blocked by D, but not lateral association.Inhibition of protofibril growth was studied in 0.5 M NaCl, pH 7.4, where lateral association is limited by the ionic strength. Stopped-flow light scattering data show a small decrease in the initial rate of polymerization and a slightly prolonged t½ ; the final intensity is less than that for a solution of long protofibrils. This result suggests that fragment D binds to the growing protofibrils with a small effect on the initial polymerization rate, but exerts its inhibitory effect by limiting the later stages of protofibril growth. Measurements of the length of the inhibited protofibrils, calculated from diffusion coefficients obtained by dynamic light scattering, confirm that polymers containing as few as 15 monomers have been obtained. Negative stain electron microscopy also shows a clear limitation of polymer growth under these conditions.Fragment D interferes with fibrin formation by directly blocking the first assembly step: bimolecular polymerization of activated fibrin monomer molecules to form protofibrils. Fragment D apparently occupies a site normally occupied by a fibrin monomer molecule, thus forming a dead-end complex which cannot undergo further assembly.
Bayesian analysis of static light scattering data for globular proteins