A Genetically Encoded Fluorescent Amino Acid

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 103; no. 26; pp. 9785 - 9789
• Main Authors: Summerer, Daniel, Chen, Shuo, Wu, Ning, Deiters, Alexander, Chin, Jason W., Schultz, Peter G.
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 27.06.2006; National Acad Sciences
• Subjects: Alanine - analogs & derivatives; Alanine - chemistry; Alanine - genetics; Alanine - metabolism; Amino acids; Amino Acids - chemistry; Amino Acids - genetics; Amino Acids - metabolism; Amino Acyl-tRNA Synthetases - genetics; Amino Acyl-tRNA Synthetases - metabolism; Biochemistry; Biological Sciences; Chemical synthesis; Codon, Nonsense - metabolism; Dansyl Compounds - chemistry; Dansyl Compounds - metabolism; Fluorescence; Fluorescent Dyes - chemistry; Genetic Code; Genetic mutation; Genetics; Humans; Plasmids; Proteins; RNA, Transfer - genetics; RNA, Transfer - metabolism; Saccharomyces cerevisiae; Saccharomyces cerevisiae - genetics; Superoxide Dismutase - chemistry; Superoxide Dismutase - metabolism; Transfer RNA; Wavelengths; Yeasts
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.0603965103

The ability to introduce fluorophores selectively into proteins provides a powerful tool to study protein structure, dynamics, localization, and biomolecular interactions both in vitro and in vivo. Here, we report a strategy for the selective and efficient biosynthetic incorporation of a low-molecular-weight fluorophore into proteins at defined sites. The fluorescent amino acid 2-amino3-(5-(dimethylamino)naphthalene-l-sulfonamide)propanoic acid (dansylalanine) was genetically encoded in Saccharomyces cerevisiae by using an amber nonsense codon and corresponding orthogonal tRNA/aminoacyl-tRNA synthetase pair. This environmentally sensitive fluorophore was selectively introduced into human superoxide dismutase and used to monitor unfolding of the protein in the presence of guanidinium chloride. The strategy described here should be applicable to a number of different fluorophores in both prokaryotic and eukaryotic organisms, and it should facilitate both biochemical and cellular studies of protein structure and function.