A straightforward protocol for the site-specific incorporation of a 19F label into any protein in vivo is described. This is done using a plasmid containing an orthogonal aminoacyl-tRNA synthetase/tRNACUA that incorporates L-4-trifluoromethylphenylalanine in response to the amber codon UAG. This method improves on other in vivo methods because the 19F 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 19F-labeled protein is similar to expressing protein in Escherichia coli and takes 4 d to obtain pure protein starting from the appropriate vectors.
|Number of pages
|Published - Oct 2007
Bibliographical noteFunding Information:
ACKNOWLEDGMENTS We thank Cinthia Kinsland for her helpful discussions on molecular biology and media. This work was supported by F&M Hackman and Eyler funds and NSF-MCB-0448297, Research Corporation (CC6364) and ACS-PRF (42214-GB4).
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology (all)