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Chemical-Tag Labeling of Proteins Using Fully Recombinant Split Inteins

Part of the Methods in Molecular Biology book series (MIMB,volume 1266)

Abstract

Chemical-tag labeling of proteins involving split inteins is an approach for the selective chemical modification of proteins without the requirement of any chemical synthesis to be performed. In a two-step protocol, a very short tag fused to a split intein auxiliary protein is first labeled in a bioconjugation reaction with a synthetic moiety either at its N-terminus (amine-tag) or at the side chain of an unnatural amino acid (click-tag). The labeled protein is then mixed with the protein of interest fused to the complementary intein fragment. In the resulting spontaneous protein trans-splicing reaction the split intein fragments remove themselves and ligate the tag to the protein of interest in a virtually traceless fashion. The reaction can be performed either using a purified protein of interest or to label a protein in the context of a living cell. All protein components are recombinantly expressed and all chemical reagents are commercially available.

Key words

  • Bioconjugation
  • Protein labeling
  • Intein
  • Protein splicing
  • Click chemistry
  • Synthetic label
  • Protein expression
  • Fluorophore

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  • DOI: 10.1007/978-1-4939-2272-7_10
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Acknowledgements

We thank Peter G. Schultz (The Scripps Research Institute) for providing plasmids for AzF incorporation. Financial support was kindly provided by the DFG (SPP1623).

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Correspondence to Henning D. Mootz .

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Bachmann, AL., Matern, J.C.J., Schütz, V., Mootz, H.D. (2015). Chemical-Tag Labeling of Proteins Using Fully Recombinant Split Inteins. In: Gautier, A., Hinner, M. (eds) Site-Specific Protein Labeling. Methods in Molecular Biology, vol 1266. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2272-7_10

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  • DOI: https://doi.org/10.1007/978-1-4939-2272-7_10

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