Modulation of a protein-folding landscape revealed by AFM-based force spectroscopy notwithstanding instrumental limitations
Devin T. Edwards, Marc-André LeBlanc, Thomas T. Perkins
Abstract
Significance Atomic force microscopy (AFM) is widely applied to unfold proteins. Using advanced cantilevers in a commercial AFM to improve stability and precision, we unfolded and then refolded an individual fast-folding, mechanically labile protein—a technically challenging sample—thousands of times. To address concerns that the measured dynamics of such proteins are dominated by the AFM assay, we modulated the protein’s folding rate using pH. Kinetic analysis demonstrated changes in free-energy landscape parameters with pH, despite the reconstructed landscape being dominated by the assay. Hence, AFM using advanced cantilevers can yield biophysical insight into proteins, even if they unfold at low force.