Direct evidence of the multidimensionality of the free-energy landscapes of proteins revealed by mechanical probes.

The study of mechanical unfolding, through the combined efforts of atomic force microscopy and simulation, is yielding fresh insights into the free-energy landscapes of proteins. Thus far, experiments have been mostly analyzed with one-dimensional models of the free-energy landscape. We show that as the two ends of a protein, filamin, are pulled apart at a speed tending to zero, the measured mechanical strength plateaus at approximately 30 pN instead of going toward zero, deviating from the Bell model. The deviation can only be explained by a switch between parallel pathways. Insightful analysis of mechanical unfolding kinetics needs to account for the multidimensionality of the free-energy landscapes of proteins, which are crucial for understanding the behavior of proteins under the small forces experienced in vivo.