Locating the Barrier for Folding of Single Molecules under an External Force

Single-molecule pulling experiments on the folding of biomolecules are usually interpreted with one-dimensional models in which the dynamics occurs on the pulling coordinate. Paradoxically, the free-energy profile along this coordinate may lack a refolding barrier, yet a barrier is known to exist for folding; thus, it has been argued that pulling experiments do not probe folding. Here, we show that transitions monitored in pulling experiments probe the true folding barrier but that the barrier may be hidden in the projection onto the pulling coordinate. However, one-dimensional theory using the pulling coordinate still yields physically meaningful energy landscape parameters.