The SNAP-tag®, CLIP-tag™, and ACP-tag/MCP-tag protein labeling technologies offer an innovative alternative to traditional fluorescent proteins for studying the function and localization of proteins in living and fixed cells. Covalent protein labeling brings simplicity and versatility to the imaging of mammalian proteins in live cells, as well as the ability to capture proteins in vitro. The creation of a single genetic construct generates a fusion protein which, when covalently attached to a variety of substrates, including fluorophores, biotin, and beads, provides a powerful tool for studying proteins. In the SNAP-tag and CLIP-tag sytems, the fusion protein is labeld by a self-labeling tag based on the DNA repair protein O6-alkylguanine-DNA-alkyltrasferase, whereas with the ACP-tag and MCP-tag systems, the labeling is catalyzed through a post-translational modification by a phosphopantetheinyl transferase.
Another way of labeling proteins is by using intein-mediated protein ligation (1) or expressed protein ligation. The IPL reaction allows the ligation of a synthetic peptide or a protein with an N-terminal cysteine residue to the thioester on the C-terminus of an expressed protein through a native peptide bond. The IPL protocol employs an IMPACT™ (NEB #E6901) C-terminal fusion vector to express and purify a protein of interest and to generate a thioester at its C-terminus. IPL allows for ligation of peptides (may contain modified amino acids) or proteins to the C-terminus of an IMPACT expressed protein.
SNAP-tag® is a registered trademark of England Biolabs, Inc.
CLIP-tag™ and IMPACT™ are trademark of New England Biolads, Inc.
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