Pressure-dependent chemical shifts in the R3 domain of Talin show that It Is thermodynamically poised for binding to either vinculin or RIAM

Talin mediates attachment of the cell to the extracellular matrix. It is targeted by the Rap1 effector RIAM to focal adhesion sites and subsequently undergoes force-induced conformational opening to recruit the actin-interacting protein vinculin. The conformational switch involves the talin R3 domain, which binds RIAM when closed and vinculin when open. Here, we apply pressure to R3 and measure (1)H, (15)N, and (13)C chemical shift changes, which are fitted using a simple model, and indicate that R3 is only 50% closed: the closed form is a four-helix bundle, while in the open state helix 1 is twisted out. Strikingly, a mutant of R3 that binds RIAM with an affinity similar to wild-type but more weakly to vinculin is shown to be 0.84 kJ mol(-1) more stable when closed. These results demonstrate that R3 is thermodynamically poised to bind either RIAM or vinculin, and thus constitutes a good mechanosensitive switch.