New Insights into Fibrin (ogen) Structure and Function

Blood clots. What? Where? When? Why? and How? These are the questions we are taught to ask and explore early in our childhood. Together we will explore the answers to these questions, limiting the scope of our discussion to the structural and functional role that fibrinogen and fibrin play in blood coagulation. History In 1859 Denis de Commercy [ 171 proposed that a soluble precursor to fibrin existed and that it could be salted out of plasma. In addition, upon denaturation it could be converted to fibrin. He called this precursor fibrinogen. Independent work by Schmidt [8l] and Hammarsten [44,45] demonstrated conclusively that a “thrombic” enzyme was required and that it might be responsible for a small amount of protein released in the conversion. In the first recorded structural experiment, Bailey et al. [ I ] placed a concentrated solution of fibrinogen (which forms a viscous strand) and a fibrin thread in an x-ray beam. The resulting patterns were indistinguishable From each other, but also from keratin and myosin. They all showed a characteristic 5.1 A repeat spacing that was later determined to be due to supercoiled a-helices [16]. Around this same time a different physiochemical “picture” of fibrinogen was emerging. Techniques including electrophoresis, ultracentrifugation, viscosity measurements and light scattering were coming into vogue and determined that fibrinogen was a prolate ellipsoid with a length between 500700 A, axial ratio between 5-20 A, and a molecular weight near 340 kDa ([I 1,3 1,78,84]). Electron microscopists Hall and Slayter [43] produced shadowcast photographs of fibrinogen and observed three, unconnected globules in a line. The center globule was the smallest, with a diameter of approximately 50 A; the terminal globules were found to be of equivalent size, approximately 60 A. The total molecule length was estimated to be 475 A; thus, the connections between the globules were approximated to 150 A. This three-domain structure has become known as the Hall and Slayter model of fibrinogen. Cohen [ 131 suggested that the globules could be linked by a-helices, which would explain the characteristic a-helix 5.1 A repeat spacing observed during fiber diffraction. A period of debate ensued because the images of the three globules could not be repeated with any of the new and more sensitive EM techniques becoming available; rather, often a single large globule was observed, prompting other models for fibrinogen [50,75]. The matter was finally resolved in 1979, when Fowler and Erickson [38] produced a definitive series of electron micrographs using both the newer negative staining and older shadow cast techniques to produce images equivalent to Hall and Slayter’s. In retrospect, the long delay in confirming Hall and Slayter’s work is due to the fact that native fibrinogen is a fragile entity that is easily denatured under many of the conditions used. The reconstruction of fibrinogen from both enzymatic and chemical fragmentation products has allowed an exact picture of the structure and function of fibrinogen to emerge. Many fragments were subjected to protein sequencing and the sequences reassembled until the whole structure was determined. These fragments also gave insights into the polymerization process as well as the shape of fibrinogen. Because plasmin is the natural protease for fibrin removal, it was the obvious choice for initial fragmentation. Nussenzweig et al. [74] characterized such a fibrinogen