Expanding the fluorous arsenal: tetrafluorinated phenylalanines for protein design

Tetrafluorinated phenylalanines were incorporated into villin headpiece subdomain HP35, and their energetic contribution to protein stability was evaluated. In comparison to pentafluoro-phenylalanines, HP35 variants harboring tetrafluorinated phenylalanines exhibit increased thermal and thermodynamic stability by a large margin. The favorable stabilization is attributed to the electrostatic ArH...pi interactions retained and strengthened by the highly, but not fully, fluorinated aromatic rings.