Acceleration of fibrin gel formation by unrelated proteins.

Addition of gamma-globulin, serum albumin, hemoglobin, or ovalbumin in concentrations of 1-10 g/dl to solutions of purified fibrinogen results in a substantial (up to six-fold) decrease in the lag time preceding appearance of a firm fibrin gel following addition of thrombin at 24 degrees C. The effect does not appear to be due to a protein-induced enhancement in the enzymatic activity of thrombin, nor does it appear to be due to the co-condensation of the added protein with fibrin/fibrinogen. It is suggested that the observed effect is primarily due to nonspecific volume exclusion arising from increased fractional occupancy of solution volume by macromolecules.

Effects of crosslinking on the rigidity and proteolytic susceptibility of human fibrin clots.

Clots formed in reconstituted human plasma or from purified human fibrin were studied in order to assess the effects of subunit crosslinking on clot strength and on resistance to plasmin degradation. The relative amounts of alpha chain and gamma chain ligation were varied by adding factor XIII to the samples. We observe, as have others, that appreciable gamma-gamma crosslinking always precedes detectable formation of alpha dimer or alpha polymer. Non-invasive light scattering measurements of the shear modulus G(t) indicate that ligation of gamma chains and of alpha chains have qualitatively similar effects on clot strength. Since alpha crosslinking occurs very slowly in the clots which are formed from plasma, we infer that under physiological conditions the involvement of alpha chains in the development of clot strength probably is only a secondary function. Light scattering techniques also were used to study the size of particles shed from the surfaces of fibrin clots undergoing fibrinolysis, and no differences could be discerned as resulting from ligation of alpha chains.

Molecular properties of lamprey fibrinogen.

Application of conventional methods not previously used for preparing lamprey fibrinogen has yielded preparations of very high purity and stability. These preparations made possible a detailed evaluation of molecular weights in normal and chaotropic buffers. The native molecule repeatedly gave a molecular weight of 352,000 to 358,000 via sedimentation equilibrium in citrate buffer; a molecular weight of 354,500 was obtained in guanidine buffer. An anomalous increase of the value of the measured apparent partial specific volume was observed in guanidine buffer. Molecular weight data of the reduced and alkylated subunits of lamprey fibrinogen, obtained by four different methods, have led us to assign molecular weights of 110,000 for (A)alpha, 72,000 for (B) beta, and 50,000 for gamma. Based upon these molecular weights obtained for the subunits, as well as that of the native fibrinogen molecule, the subunit composition can best be fitted to the formulation [(A)n alpha, (B) beta 2, gamma 2] rather than the conventional[(A) alpha 2, (B) beta 2, gamma 2] which would yield a molecular weight of 464,000. Analysis of a stabilized clot induced by Ca2+ showed only gamma dimers; alpha subunit polymerization was undetectable. Cross-linking of lamprey fibrin in the presence of dansylcadaverine and Ca2+ results in fluorescent labeling of the gamma chains and to a lesser extent the gamma dimer. Differing from other reported vertebrate cross-linking systems, the lamprey fibrin alpha subunit appears essentially unreactive in both polymer formation and dansylcadaverine incorporation. These distinct molecular properties may be reasonably attributed to the existence of the single (A)n alpha subunit in the molecular structure of the molecule.