Chemistry & Biology
Volume 15, Issue 2, 22 February 2008, Pages 128-136
Journal home page for Chemistry & Biology

Article
An Engineered Protein Tag for Multiprotein Labeling in Living Cells

https://doi.org/10.1016/j.chembiol.2008.01.007Get rights and content
Under an Elsevier user license
open archive

Summary

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.

CHEMBIO

Cited by (0)

3

Present address: Pasteur Institute, Unité d'Immunologie Structurale, 25–28 rue du Docteur Roux, 75724 Paris Cedex 15, France.

4

Present address: Division of Protein and Nucleic Acid Chemistry, MRC Laboratory of Molecular Biology, Hills Road, Cambridge CB2 2QH, United Kingdom.

5

Present address: Sanofi-Aventis Deutschland, Chemical Sciences, D-65926 Frankfurt am Main, Germany.