Stabilization of coiled-coil peptide domains by introduction of trifluoroleucine

Substitution of leucine residues by 5,5,5-trifluoroleucine at the d-positio ns of the leucine zipper peptide GCN4-pld increases the thermal stability o f the coiled-coil structure. The midpoint thermal unfolding temperature of the fluorinated peptide is elevated by 13 degreesC at 30 muM peptide concen tration. The modified peptide is more resistant to chaotropic denaturants, and the free energy of folding of the fluorinated peptide is 0.5-1.2 kcal/m ol larger than that of the hydrogenated form. A similarly fluorinated form of the DNA-binding peptide GCN4-bZip binds to target DNA sequences with aff inity and specificity identical to those of the hydrogenated form, while de monstrating enhanced thermal stability. Molecular dynamics simulation on th e fluorinated GCN4-pld peptide using the Surface Generalized Born implicit solvation model revealed that the coiled-coil binding energy is 55% more fa vorable upon fluorination. These results suggest that fluorination of hydro phobic substructures in peptides and proteins may provide new means of incr easing protein stability, enhancing protein assembly, and strengthening rec eptor-ligand interactions.