Computational Modeling of the Thermodynamics of the Mesophilic and Thermophilic Mutants of Trp-Cage Miniprotein

[Image: see text] We characterize the folding–unfolding thermodynamics of two mutants of the miniprotein Trp-cage by combining extended molecular dynamics simulations and an advanced statistical–mechanical-based approach. From a set of molecular dynamics simulations in an explicit solvent performed along a reference isobar, we evaluated the structural and thermodynamic behaviors of a mesophilic and a thermophilic mutant of the Trp-cage and their temperature dependence. In the case of the thermophilic mutant, computational data confirm that our theoretical–computational approach is able to reproduce the available experimental estimate with rather good accuracy. On the other hand, the mesophilic mutant does not show a clear two-state (folded and unfolded) behavior, preventing us from reconstructing its thermodynamics; thus, an analysis of its structural behavior along a reference isobar is presented. Our results show that an extended sampling of these kinds of systems coupled to an advanced statistical–mechanical-based treatment of the data can provide an accurate description of the folding–unfolding thermodynamics along a reference isobar, rationalizing the discrepancies between the simulated and experimental systems.