Effect of Main and Side Chains on the Folding Mechanism of the Trp-Cage Miniprotein.

Proteins that do not fold into their functional native state have been linked to diseases. In this study, the influence of the main and side chains of individual amino acids on the folding of the tryptophan cage (Trp-cage), a designed 20-residue miniprotein, was analyzed. For this purpose, we calculated the solvation free energy (SFE) contributions of individual atoms by using the 3D-reference interaction site model with the atomic decomposition method. The mechanism by which the Trp-cage is stabilized during the folding process was examined by calculating the total energy, which is the sum of the conformational energy and SFE. The folding process of the Trp-cage resulted in a stable native state, with a total energy that was 62.4 kcal/mol lower than that of the unfolded state. The solvation entropy, which is considered to be responsible for the hydrophobic effect, contributed 31.3 kcal/mol to structural stabilization. In other words, the contribution of the solvation entropy accounted for approximately half of the total contribution to Trp-cage folding. The hydrophobic core centered on Trp6 contributed 15.6 kcal/mol to the total energy, whereas the solvation entropy contribution was 6.3 kcal/mol. The salt bridge formed by the hydrophilic side chains of Asp9 and Arg16 contributed 10.9 and 5.0 kcal/mol, respectively. This indicates that not only the hydrophobic core but also the salt bridge of the hydrophilic side chains gain solvation entropy and contribute to stabilizing the native structure of the Trp-cage.