A molecular dynamics simulation study of the unfolding of barnase induced by reaction field perturbation

We present a novel method to study protein unfolding by molecular dynamics (MD) simulations. A term resembling a reaction field is added as a perturbation to the Ewald sum, the most commonly used method to calculate electrostatic interactions in MD simulations. This reaction field perturbation (RFP) mimics qualitatively on the level of molecular interactions the changes in a protein solvent system when denaturants (e.g., urea or guadinium chloride) are added. The method is tunable by two parameters that control strength and “type’' (i.e., different regions of the electrostatic interactions can be weakened selectively) of the perturbations. The approach is tested by a detailed unfolding study of barnase based on several simulations, starting from two initial structures and with varying strength of the RFP. Unfolding is studied by following the changes in the radius of gyration, in the root mean square deviation and in the solvent accessible surface area of the protein. On the local level, the loss of secondary structure is monitored. In addition, the calculation of Voronoi volumes of individual side chains and the number of intruding waters as a function of simulation time are used to analyze the hydrophobic cores. We find an unfolding pathway and folding intermediates which agree well with the experimental data available and which are consistent with earlier simulation studies. Since under one set of RFP conditions the intermediate states are stable for approximately 100 ps, it is possible to characterize these states resembling the experimentally observed transition state and folding intermediate in great detail.