From folding theories to folding proteins: A review and assessment of simulation studies of protein folding and unfolding

Beginning with simplified lattice and continuum "minimalist" models and pro gressing to detailed atomic models, simulation studies have augmented and d irected development of the modern landscape perspective of protein folding. In this review we discuss aspects of detailed atomic simulation methods ap plied to studies of protein folding free energy surfaces, using biased-samp ling free energy methods and temperature-induced protein unfolding. We revi ew studies from each on systems of particular experimental interest and ass ess the strengths and weaknesses of each approach in the context of "exact" results for both free energies and kinetics of a minimalist model for a be ta-barrel protein. We illustrate in detail how each approach is implemented and discuss analysis methods that have been developed as components of the se studies. We describe key insights into the relationship between protein topology and the folding mechanism emerging from folding free energy surfac e calculations. We further describe the determination of detailed "pathways " and models of folding transition states that have resulted from unfolding studies. Our assessment of the two methods suggests that both can provide, often complementary, details of folding mechanism and thermodynamics, but this success relies on (a) adequate sampling of diverse conformational regi ons for the biased-sampling free energy approach and (b) many trajectories at multiple temperatures for unfolding studies. Furthermore, we find that t emperature-induced unfolding provides representatives of folding trajectori es only when the topology and sequence (energy) provide a relatively funnel ed landscape and "off-pathway" intermediates do not exist.