Molecular dynamics simulations reveal how vinculin refolds partially unfolded talin rod helices to stabilize them against mechanical force

Vinculin binds to specific sites of mechanically unfolded talin rod domains to reinforce the coupling of the cell's exterior to its force generation machinery. Force-dependent vinculin-talin complexation and dissociation was previously observed as contraction or extension of the unfolded talin domains respectively using magnetic tweezers. However, the structural mechanism underlying vinculin recognition of unfolded vinculin binding sites (VBSs) in talin remains unknown. Using molecular dynamics simulations, we demonstrate that a VBS dynamically refolds under force, and that vinculin can recognize and bind to partially unfolded VBS states. Vinculin binding enables refolding of the mechanically strained VBS and stabilizes its folded alpha-helical conformation, providing resistance against mechanical stress. Together, these results provide an understanding of a recognition mechanism of proteins unfolded by force and insight into the initial moments of how vinculin binds unfolded talin rod domains during the assembly of this mechanosensing meshwork.Cells attach to and interact with their surroundings, and sense mechanical signals from the microenvironment. Vinculin and talin are classical examples of mechanosensitive proteins that complex in a force-dependent manner. Talin domains unfold under mechanical loads exposing buried vinculin binding sites, namely single alpha-helices. Such mechanical stretching enables recognition by vinculin, however, the structural states of vinculin's binding site when under force and the complexation mechanisms are poorly understood. We used molecular dynamics simulations to study the behavior of a single vinculin binding helix under mechanical load and during the initial moments of the vinculin-talin binding process. Our results reveal the highly dynamic nature of a vinculin binding helix under mechanical load. Depending on the force magnitude, the helix frequently reforms to a partially folded conformation on the nanosecond to microsecond timescale. Crucially, vinculin recognizes such partially folded helices in talin and promotes their refolding against force. Our findings are beyond the resolution limits imposed by experimental single molecule methods, suggesting a model which may be useful as a general mechanism for protein-protein binding under mechanical load.