Analysis of the Free-Energy Surface of Proteins from Reversible Folding Simulations

Computer generated trajectories can, in principle, reveal the folding pathways of a protein at atomic resolution and possibly suggest general and simple rules for predicting the folded structure of a given sequence. While such reversible folding trajectories can only be determined ab initio using all-atom transferable force-fields for a few small proteins, they can be determined for a large number of proteins using coarse-grained and structure-based force-fields, in which a known folded structure is by construction the absolute energy and free-energy minimum. Here we use a model of the fast folding helical λ-repressor protein to generate trajectories in which native and non-native states are in equilibrium and transitions are accurately sampled. Yet, representation of the free-energy surface, which underlies the thermodynamic and dynamic properties of the protein model, from such a trajectory remains a challenge. Projections over one or a small number of arbitrarily chosen progress variables often hide the most important features of such surfaces. The results unequivocally show that an unprojected representation of the free-energy surface provides important and unbiased information and allows a simple and meaningful description of many-dimensional, heterogeneous trajectories, providing new insight into the possible mechanisms of fast-folding proteins.Author Summary: The process of protein folding is a complex transition from a disordered to an ordered state. Here, we simulate a specific fast-folding protein at the point at which the native and denatured states are at equilibrium and show that obtaining an accurate description of the mechanisms of folding and unfolding is far from trivial. Using simple quantities which quantify the degree of native order is, in the case of this protein, clearly misleading. We show that an unbiased representation of the free-energy surface can be obtained; using such a representation we are able to redesign the landscape and thus modify, upon site-specific “mutations”, the folding and unfolding rates. This leads us to formulate a hypothesis to explain the very fast folding of many proteins.