Monte Carlo simulation of mechanical unfolding of proteins based on a simple two-state model

Single molecule methods are becoming routine biophysical techniques for studying biological macromolecules. In mechanical unfolding of proteins, an externally applied force is used to induce the unfolding of individual protein molecules. Such experiments have revealed novel information that has significantly enhanced our understanding of the function and folding mechanisms of several types of proteins. To obtain information on the unfolding kinetics and the free energy landscape of the protein molecule from mechanical unfolding data, a Monte Carlo simulation based on a simple two-state kinetic model is often used. In this paper, we provide a detailed description of the procedure to perform such simulations and discuss the approximations and assumptions involved. We show that the appearance of the force versus extension curves from mechanical unfolding of proteins is affected by a variety of experimental parameters, such as the length of the protein polymer and the force constant of the cantilever. We also analyze the errors associated with different methods of data pooling and present a quantitative measure of how well the simulation results fit experimental data. These findings will be helpful in experimental design, artifact identification, and data analysis for single molecule studies of various proteins using the mechanical unfolding method.