Preparation of site-specifically labeled fluorinated proteins for 19F-NMR structural characterization

A straightforward protocol for the site-specific incorporation of a F-19 label into any protein in vivo is described. This is done using a plasmid containing an orthogonal aminoacyl-tRNA synthetase/tRNA(CUA) that incorporates L-4-trifluoromethylphenylalanine in response to the amber codon UAG. This method improves on other in vivo methods because the F-19 label is incorporated into only one location on the protein of interest and that protein can easily be produced in large quantities at low cost. The protocol for producing F-19-labeled protein is similar to expressing protein in Escherichia coli and takes 4 d to obtain pure protein starting from the appropriate vectors.