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Polyfluorinated N-alpha-Fmoc-is an element of-Boc-L-lysine represents the best-in-class among a set of polyfluorinated amino acids (PFAs) which are useful tools for (19)F NMR-Based Screening. In this communication, optimized reaction conditions that allowed for the multi-gram preparation of this unnatural amino acid are reported.

Two novel series of polyfluorinated amino acids (PFAs) were designed and synthesized according to a very short and scalable synthetic sequence. The advantages and limitations of these moieties for screening purposes are presented and discussed. The potential applications of these PFAs were tested with their incorporation into small arginine-containing peptides that represent suitable substrates for the enzyme trypsin. The enzymatic reactions were monitored by 19F NMR spectroscopy, using the 3-FABS (three fluorine atoms for biochemical screening) technique. The high sensitivity achieved with these PFAs permits a reduction in substrate concentration required for 3-FABS. This is relevant in the utilization of 3-FABS in fragment-based screening for identification of small scaffolds that bind weakly to the receptor of interest. The large dispersion of 19F isotropic chemical shifts allows the simultaneous measurement of the efficiency of the different substrates, thus identifying the best substrate for screening purposes. Furthermore, the knowledge of KM and Kcat for the different substrates allows the identification of the structural motifs responsible for the binding affinity to the receptor and those affecting the chemical steps in enzymatic catalysis. This enables the construction of suitable pharmacophores that can be used for designing nonpeptidic inhibitors with high affinity for the enzyme or molecules that mimic the transition state. The novel PFAs can also find useful application in the FAXS (fluorine chemical shift anisotropy and exchange for screening) experiment, a 19F-based competition binding assay for the detection of molecules that inhibit the interaction between two proteins.