Oxidation Effects on the von Willebrand Factor A2 Domain Investigated by Molecular Dynamics Simulations

Abstract

Inflammation due to traumatic injury causes the release of oxidants that alter the function of blood coagulation factors leading to a potential pro-thrombotic state. Studying the effects of oxidative stress onto the blood coagulation process is essential to develop treatments for trauma patients. Recent experimental evidence has shown that in the presence of hypochlorous acid (HOCl), the protein von Willebrand Factor (VWF) is more active in tethering platelets and much harder to get cleaved by the metalloprotease ADAMTS13. These observations were linked to the increased oxidation of methionine residues in the A1, A2 and A3 domains of VWF with increasing concentrations of HOCl. The exact mechanism how oxidation of methionine residues causes an increase in VWF activity is currently not understood at the molecular level. In the present work, we used steered molecular dynamics simulations and free energy perturbation methods to study how oxidation of methionine residues alters the kinetic and thermodynamic stability of the A2 domain. Previous studies have shown that the A2 domain, which contains the cleavage site for ADAMTS13, is sensitive to tensile force. The results presented here show that methionine oxidation makes the C-terminal helix of the A2 domain detach from the rest of the protein at a lower force, thus accelerating unfolding. Furthermore, oxidation of methionine residues increases the likelihood of finding the protein in the denatured state. It is plausible that unfolding of the A2 domain favors exposure of its neighboring A1 domain, which contains the binding site to platelets, thus activating VWF. This study highlights how the conversion from methionine to methionine sulfoxide, making the residue hydrophilic, destabilizes the native state leading to a modified biological pathway.