Buffers strongly modulate fibrin self-assembly into fibrous networks.
ArticleKurniawan, N.A., van Kempen, T.H.S., Sonneveld, S., Rosalina, T., Vos, B., Jansen, K., Peters, G.W.M. & van de Vosse, F.N. (2017). Buffers strongly modulate fibrin self-assembly into fibrous networks. Langmuir, 33(25), 6342-6352. In Scopus Cited 0 times.
Fibrin is a plasma protein with a central role in blood clotting and wound repair.
Upon vascular injury, fibrin forms resilient fibrillar networks (clots) via a multi-step self assembly process, from monomers, to double-stranded protofibrils, to a branched network of thick fibers. In vitro, fibrin self-assembly is sensitive to physicochemical conditions like the solution pH and ionic strength, which tune the strength of the non-covalent driving forces. Here we report a surprising finding that the buffer—which is necessary to control the pH and is typically considered to be inert—also significantly influences fibrin self-assembly. We show by confocal microscopy and quantitative light-scattering that various common buffering agents have no effect on the initial assembly of fibrin monomers into protofibrils but strongly hamper the subsequent lateral association of protofibrils into thicker fibers. At 3 mg/ml fibrin, the average
number of protofibrils per fiber drops nearly 10-fold, from 180 in the presence of 20 mM HEPES to only 20 in the presence of 200 mM HEPES. By using different enzymes to initiate fibrin self assembly, we reveal that the mechanism of buffer-induced changes in fibrin assembly involves weakening of the interactions between the so-called αC-region, which are long flexible appendages on fibrin that couple adjacent protofibrils. We further find that the structural changes are independent of the molecular structure of the buffering agents and even occur in fibrin networks formed from platelet-poor plasma, suggesting that buffers act by changing the hydrogen bonding network structure of the hydration layer surrounding fibrin. This buffer mediated decrease in protofibril bundling results in a marked reduction in the permeability of fibrin networks. Yet, the buffers only weakly influence the elastic modulus of fibrin networks
because the effect of higher fiber density is cancelled out by the effect of the smaller stiffness of thinner fibers. Buffers can therefore be used as a tuning parameter to independently control the elastic properties and the permeability of fibrin networks, which is useful in studies of cell adhesion and migration. Our work raises the possibility that fibrin assembly in vivo may be regulated by variations in the acute-phase levels of bicarbonate and phosphate, which act as physiological buffering agents of blood pH. Moreover, our findings add a striking new example of buffer-induced effects on biomolecular self-assembly to recent findings for a range of proteins and lipids.