PB0531 - Mechanical behavior of clots: Molecular mechanism of forced unfolding of crosslinked fibrin protofibrils

Background: Fibrin is the scaffold of blood clots and thrombi providing them with the mechanical stability necessary to withstand deformations and rupture. The mechanical behavior of cross-linked fibrin clots is largely determined by stretching of fibers and protofibrils followed by forced molecular unfolding, which is not well understood. Owing to the structural complexity of fibrin protofibrils, the unfolding molecular transitions in fibrin must be studied by a combination of stretching experiments and biomolecular simulations to provide detailed structure-based interpretations of the experimental data.

Aims: To investigate the mechanical properties of fibrin protofibrils and to resolve the structural bases underlying the mechanisms of their forced elongation.

Methods: Using atomic force microscopy (AFM), we pulled upward individual fibrin protofibrils formed on a mica surface. The resulting saw-tooth-like force-extension curves displayed force peaks corresponding to elementary unfolding transitions of compact molecular structures. To help interpret the experimental molecular unfolding profiles, we employed Langevin simulations and the coarse-grained Self Organized Polymer model to complement experiments with dynamic force measurements in silico on a short fibrin protofibril fragment composed of four fibrin monomers in the upper and lower strand (FP4/4). The force-extension profiles from AFM-based single-protofibril unfolding experiments in vitro and in silico were directly correlated.

Results: The experimental and theoretical histograms of peak forces revealed similar most probable unfolding forces, i.e. 230 pN (experiment) versus 220 pN (simulations), which validates our combined experimental and theoretical approach (Figure). We identified four most important molecular transition types, which include the disruption of D:D interfaces and stepwise unfolding of the γ nodules (Table).

Conclusion(s): We illuminated structural mechanisms and mapped the free-energy landscape for forced unfolding of fibrin protofibrils. This information is important to understand the basis of fibrin mechanics, and it has far reaching implications for the deformability and integrity of fibrin clots and thrombi.