Stabilizing and Destabilizing Effects of Phenylalanine → F₅-Phenylalanine Mutations on the Folding of a Small Protein

We report a systematic evaluation of phenylalanine-to-pentafluorophenylalanine (Phe → F₅-Phe) mutants for the 35-residue chicken villin headpiece subdomain (c-VHP), the hydrophobic core of which features a cluster of three Phe side chains (residues 6, 10, and 17). Phe → F₅-Phe mutations are interesting because aryl−perfluoroaryl interactions of optimal geometry are intrinsically more favorable than aryl−aryl interactions and because perfluoroaryl units are more hydrophobic than are analogous aryl units. One mutant, Phe-10 → F₅-Phe, provides enhanced tertiary structural stability relative to the native sequence. The other six mutants analyzed caused a decrease in stability.