Fig. 5.
Typical folding trajectories for Q (the fraction of native contact formation), pseudo Q (the fraction of native single-water separated contact formation), and contact energies as a function of integration time steps, providing molecular details of kinetic traps controlling folding of V44T and V53T. Folding simulations of V44T are shown in A and B, and those of V53T are shown in C and D. Both calculations are carried out at 0.9 Tf, where Tf is the folding temperature shown at Table 3 (Supporting Text). In the snapshots at both B and D (where contact energies are plotted as a function of time steps), blue spheres are used to identify single-water separated contacts (notice that unfolded residues without blue spheres are fully solvated). We color residues at native conformations in red to identify folded regions and the unstructured residues in gray. (B) For V44T, position 44 is colored purple for visual guidance, and snapshot (a) shows a typical unfolded configuration where short-range contacts are formed. Snapshot b shows a typical kinetic trap where the removal of favorable hydrophobic contacts at this position sabotages nascent native contact formation at the β2, β3, and β4 regions. These β sheets are frayed, and their pairing distances to position 44 are beyond the size of a single water (i.e., fully solvated, highlighted in a cyan box). As a consequence, the distribution of contact formation shifts toward the RT loop at the early stage of folding; an alignment for structural-search collapse is impeded, and kinetic traps are formed. Snapshots c and d correspond to the structural-search collapse where abundant water molecules are expelled from β2-, β3-, and β4-strands, and the diverging turn. Afterward, position 44 is neatly packed at the center of the aforementioned β sheets. Snapshots e and f depict the final stage of folding where residual water molecules are expelled from the hydrophobic core as terminal β-strands pack against the rest of the protein. (D) Next we discuss the folding event of V53T mutant. Position 53 is colored in green to provide visual guidance. Snapshot a is a typical trap state, which shows a collapsed yet seemingly flat conformation with short-range contacts mostly formed at the turn and loop regions. A cyan box highlights position 53, which is exposed to a fully solvated yet unformed hydrophobic core. Snapshot b shows a typical unfolded state where position 53 is fully solvated. Snapshots (c and d) indicate a structural-search collapse in which position 53 is exposed to the hydrated hydrophobic core (as illustrated by a cyan box) where water molecules are found between β4 and the RT loop. At the last stage of folding, the desolvation of hydrophobic core takes place; water molecules are expelled from this core, and V53T folds to its native state (e). Simulations suggest that these two positions play distinct topological roles, accounting for the decreases of folding rates observed experimentally.