(525f) Improved Analysis of Single-Molecule Force-Spectroscopy Data for Extracting Free Energy Barrier Heights and Barrier Crossing Rates

Applying controlled amount of forces to single molecules like DNA, RNA, and proteins by means of micromanipulation techniques such as optical tweezers and AFM can provide significant information about the underlying energy landscape. These experiments are generally performed at fixed or constantly increasing forces and the dynamic response of the molecule is recorded in the form of molecular extension vs. force and extension vs. time plots. This "nonequilibrium" response may then be analyzed to extract "equilibrium" properties such as the intrinsic free energy barrier (&Delta G^*), barrier distance (x^*) and transition rate (k₀) of the molecule undergoing conformational transition between different macroscopic states. Recently, Dudko et al. (PRL, 2006; PNAS, 2008) have developed a sophisticated approach based on Kramer's theory of barrier crossing to extract the free energy barrier heights and locations as well as the rates of barrier crossing from force-spectroscopy data in which the molecule is stretched at a constant velocity. Here we present an extension of this approach that provides more accurate estimates of free energies and kinetic rates by additionally exploiting the compliance of the device itself. We also present a validation of this approach by studying a simplified system composed of a Brownian particle in a double-well potential. We are now applying this approach to determine the free energy profiles and kinetics of conformational transitions in coarse-grained models of proteins and RNA.