Synthesis of fluorinated leucines, valines and alanines for use in protein NMR.

Efficient syntheses of fluorinated leucines, valines and alanines are described. The synthetic routes provide expedient access to various C/N/D isotopologues requiring solely readily available and inexpensive isotope containing reagents such as NaBD, carbon-C dioxide and sodium azide-1-N. The lightly fluorinated leucines and valines were found to be good substrates for cell-free protein expression and even 3-fluoroalanine, which is highly toxic to bacteria , could be incorporated into proteins this way. F-NMR spectra of the protein GB1 produced with these amino acids showed large chemical shift dispersions. Particularly high incorporation yields and clean F-NMR spectra were obtained for GB1 produced with valine residues, which had been synthesized with a single fluorine substituting a hydrogen stereospecifically in one of the methyl groups.