Substitution of the Human αC Region with the Analogous Chicken Domain Generates a Fibrinogen with Severely Impaired Lateral Aggregation: Fibrin Monomers Assemble into Protofibrils but Protofibrils Do Not Assemble into Fibers

Abstract

Fibrin polymerization occurs in two steps: the assembly of fibrin monomers into protofibrils and the lateral aggregation of protofibrils into fibers. Here we describe a novel fibrinogen that apparently impairs only lateral aggregation. This variant is a hybrid, where the human αC region has been replaced with the homologous chicken region. Several experiments indicate this hybrid human-chicken (HC) fibrinogen has an overall structure similar to normal. Thrombin-catalyzed fibrinopeptide release from HC fibrinogen was normal. Plasmin digests of HC fibrinogen produced fragments that were similar to normal D and E; further, as with normal fibrinogen, the knob ‘A’ peptide, GPRP, reversed the plasmin cleavage associated with addition of EDTA. Dynamic light scattering and turbidity studies with HC fibrinogen showed polymerization was not normal. Whereas early small increases in hydrodynamic radius and absorbance paralleled the increases seen during the assembly of normal protofibrils, HC fibrinogen showed no dramatic increase in scattering as observed with normal lateral aggregation. To determine whether HC and normal fibrinogen could form a copolymer, we examined mixtures of these. Polymerization of normal fibrinogen was markedly changed by HC fibrinogen, as expected for mixed polymers. When the mixture contained 0.45 μM normal and 0.15 M HC fibrinogen, the initiation of lateral aggregation was delayed and the final fiber size was reduced relative to normal fibrinogen at 0.45 μM. Considered altogether our data suggest that HC fibrin monomers can assemble into protofibrils or protofibril-like structures but these either cannot assemble into fibers or assemble into very thin fibers.

Post-translational oxidative modification of fibrinogen is associated with coagulopathy after traumatic injury

“…Dissociation and interaction of fibrinogen αC domains from the E region enables fibrin protofibril lateral aggregation during clot formation [36-38]. Alpha C double hairpin structures interact with adjacent and complementary hairpin structures on adjacent fibrin monomers to form large beta sheet structures, eventually coalescing into fibers [39].…”

Identification of Respective Lysine Donor and Glutamine Acceptor Sites Involved in Factor XIIIa-catalyzed Fibrin α Chain Cross-linking

“…Studies indicate that in fibrinogen two ␣C domains form a dimer through interaction with each other and the central region of the molecule. In polymeric fibrin, factor XIIIa covalently cross-links ␣C regions from different fibrin molecules to produce the fibrin network (41)(42)(43). It is speculated that the Gln acceptors located near the amino terminus of fibrin ␣C regions (␣C-connector) form cross-links with Lys donors located near the carboxyl terminus of ␣C regions (␣C domain) from another fibrin molecule (44).…”

Fibrin Formation, Structure and Properties

“…Fibrinogen lacking the αC regions forms clots with thinner fibers and a higher density of branch points than clots made of full-length fibrinogen (Collet et al 2005). Furthermore, a recombinant fibrinogen construct that has the longer human αC regions replaced with the shorter chicken αC regions displayed impaired lateral aggregation of protofibrils (Ping et al 2011). …”

“…The role of the αC polymers in fibrin mechanics, underestimated in the past, has been studied intensively more recently (Houser et al 2010; Falvo et al 2010; Ping et al 2011; Helms et al 2012; Averett et al 2012). It has been shown that the transglutaminase-catalyzed covalent crosslinking of the α chains contributes substantially to the fibrin clot stiffness and elasticity (Collet et al 1999; Piechocka et al 2010; Helms et al 2012; Duval et al 2014; Kurniawan et al 2014), including the fast elastic recoil of stretched fibrin fibers (Hudson et al 2013).…”