RepTAGs: universal tags for isolation and labeling of proteins, for labeling live mammalian cells and for drug discovery.
Methods for specific immobilization, isolation and labeling of proteins are central to the elucidation of cellular functions. Based on bacterial repressor proteins, which bind to specific target sequences in response to small molecules (macrolide and tetracycline antibiotics) or environmental parameters (temperature), we have developed a set of protein tags (RepTAGs), which enable reversible immobilization of the protein of interest on a solid support for the isolation and quantification as well as for the specific labeling of target proteins with fluorescent dyes for tracking them within a complex protein mixture. Similarly, live mammalian cells were specifically labeled with a fluorescent operator sequence bound to RepTAGs, which were directed towards the cell surface for easy discrimination between transfected and untransfected cell populations. Based on the drug-responsive RepTAG-DNA interactions, it was also possible to quantify or discover antibiotics in environmental samples or compound libraries by means of rapid, sensitive detection methods involving fluorescence polarization and bioluminescence. We believe that the universally applicable RepTAGs will become essential for the analysis and manipulation of proteins in the most diverse areas of protein chemistry and cell biology.
Weber
Wilfried
W
Institute for Chemical and Bioengineering, ETH Zurich, Wolfgang-Pauli-Strasse 10, HCI F115, CH-8093 Zurich, Switzerland.
Link
Nils
N
Aubel
Dominique
D
Weber
Cornelia C
CC
Fussenegger
Martin
M
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002846:Chromatography, Affinity / Q000379:methods; D015195:Drug Design; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D004926:Escherichia coli / Q000235:genetics; D005456:Fluorescent Dyes / Q000737:chemistry; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D007668:Kidney / Q000166:cytology; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification*
D005456:Fluorescent Dyes; D011993:Recombinant Fusion Proteins