title: Simultaneous triggering of protein activity and fluorescence.
abstract: Many areas of biology can benefit greatly from methods to spatially and temporally control protein activity. Here, we describe an approach that allows the simultaneous photo-triggering of the activity and the fluorescence of a protein. Smad2, a protein central to the transforming growth factor-beta (TGF-beta) signal transduction pathway, was modified with a fluorophore and a photocleavable moiety that acted as both a caging and a fluorescence quenching group. In its caged state, the protein formed a non-fluorescent heterodimer with the protein SARA. Irradiation with UV light and photocleavage of the caging group produced a fluorescent homotrimer. These in vitro experiments demonstrated that a photochemical trigger mimicking the critical biochemical event of serine phosphorylation involved in the TGF-beta signaling pathway could be obtained and that fluorescence could be used as a read-out of protein activity. This approach should prove particularly useful for the monitoring of a protein's activity and location inside of living cells.
authors.affiliation: Laboratory of Synthetic Protein Chemistry, The Rockefeller University, New York, New York 10021, USA.
authors.forename: Jean-Philippe
authors.initials: JP
authors.lastname: Pellois
authors.forename: Michael E
authors.initials: ME
authors.lastname: Hahn
authors.forename: Tom W
authors.initials: TW
authors.lastname: Muir
mesh_terms: D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D004268:DNA-Binding Proteins / Q000032:analysis* / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism*; D005453:Fluorescence; D015394:Molecular Structure; D051899:Smad2 Protein; D013050:Spectrometry, Fluorescence; D015534:Trans-Activators / Q000032:analysis* / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism*
chemical_list: D004268:DNA-Binding Proteins; D051899:Smad2 Protein; D015534:Trans-Activators