Genetic Encoding of para-Pentafluorosulfanyl Phenylalanine: A Highly Hydrophobic and Strongly Electronegative Group for Stable Protein Interactions

SF5Phe, para-pentafluorosulfanyl phenylalanine, is an unnatural amino acid with extreme physicochemical properties, which is stable in physiological conditions. Here we present newly developed aminoacyl-tRNA synthetases that enable genetic encoding of SF5Phe for site-specific incorporation into proteins in high yields. Owing to the SF5 moiety's dichotomy of strong polarity and high hydrophobicity, the unnatural amino acid forms specific and strong interactions in proteins. The potential of SF5Phe in protein research is illustrated by (i) increasing the binding affinity of a consensus pentapeptide motif toward the β subunit of Escherichia coli DNA polymerase III holoenzyme by mutation of a phenylalanine to a SF5Phe residue, (ii) site-specifically adhering β-cyclodextrin to the surface of ubiquitin, and (iii) selective detection of 19F-19F nuclear Overhauser effects in the Escherichia coli peptidyl-prolyl cis/trans-isomerase B following mutation of two phenylalanine residues in the core of the protein to SF5Phe. With increasing use of the SF5 moiety in pharmaceutical chemistry, this general method of functionalizing proteins with SF5 groups opens unique opportunities for structural biology and in vivo studies.