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, 15 (2), 128-36

An Engineered Protein Tag for Multiprotein Labeling in Living Cells

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An Engineered Protein Tag for Multiprotein Labeling in Living Cells

Arnaud Gautier et al. Chem Biol.

Abstract

The visualization of complex cellular processes involving multiple proteins requires the use of spectroscopically distinguishable fluorescent reporters. We have previously introduced the SNAP-tag as a general tool for the specific labeling of SNAP-tag fusion proteins in living cells. The SNAP-tag is derived from the human DNA repair protein O6-alkylguanine-DNA alkyltransferase (AGT) and can be covalently labeled in living cells using O6-benzylguanine derivatives bearing a chemical probe. Here we report the generation of an AGT-based tag, named CLIP-tag, which reacts specifically with O2-benzylcytosine derivatives. Because SNAP-tag and CLIP-tag possess orthogonal substrate specificities, SNAP and CLIP fusion proteins can be labeled simultaneously and specifically with different molecular probes in living cells. We furthermore show simultaneous pulse-chase experiments to visualize different generations of two different proteins in one sample.

Comment in

  • Simultaneous Protein Tagging in Two Colors
    C Schultz et al. Chem Biol 15 (2), 91-2. PMID 18291310.
    The fluorescent tagging of proteins in the natural environment of the cell is an emerging technique in cell biology. In this issue of Chemistry & Biology, Gautier et …

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