Open AccessSite‐Specific Protein Labeling with SNAP‐Tags
Author(s) -
Cole Nelson B.
Publication year - 2013
Publication title -
current protocols in protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.409
H-Index - 32
eISSN - 1934-3663
pISSN - 1934-3655
DOI - 10.1002/0471140864.ps3001s73
Subject(s) - snap , chemistry , biology , biophysics , computational biology , computer science , computer graphics (images)
Site‐specific labeling of cellular proteins with chemical probes is a powerful tool for studying protein function in living cells. A number of small peptide and protein tags have been developed that can be labeled with synthetic probes with high efficiencies and specificities and provide flexibility not available with fluorescent proteins. The SNAP‐tag is a modified form of the DNA repair enzyme human O 6 ‐alkylguanine‐DNA‐alkyltransferase, and undergoes a self‐labeling reaction to form a covalent bond with O 6 ‐benzylguanine (BG) derivatives. BG can be modified with a wide variety of fluorophores and other reporter compounds, generally without affecting the reaction with the SNAP‐tag. In this unit, basic strategies for labeling SNAP‐tag fusion proteins, both for live cell imaging and for in vitro analysis, are described. This includes a description of a releasable SNAP‐tag probe that allows the user to chemically cleave the fluorophore from the labeled SNAP‐tag fusion. In vitro labeling of purified SNAP‐tag fusions is briefly described. Curr. Protoc. Protein Sci . 73:30.1.1‐30.1.16. © 2013 by John Wiley & Sons, Inc.