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ACS Chem Biol. 2016 Dec 16;11(12):3245-3250. Epub 2016 Oct 17.

Click-MS: Tagless Protein Enrichment Using Bioorthogonal Chemistry for Quantitative Proteomics.

Author information

1
Department of Molecular Biology, Faculty of Science, Radboud Institute for Molecular Life Sciences, Radboud University Nijmegen , Nijmegen, The Netherlands.
2
Department of Bio-organic Chemistry, Faculty of Science, Institute for Molecules and Materials, Radboud University Nijmegen , Nijmegen, The Netherlands.

Abstract

Epitope-tagging is an effective tool to facilitate protein enrichment from crude cell extracts. Traditionally, N- or C-terminal fused tags are employed, which, however, can perturb protein function. Unnatural amino acids (UAAs) harboring small reactive handles can be site-specifically incorporated into proteins, thus serving as a potential alternative for conventional protein tags. Here, we introduce Click-MS, which combines the power of site-specific UAA incorporation, bioorthogonal chemistry, and quantitative mass spectrometry-based proteomics to specifically enrich a single protein of interest from crude mammalian cell extracts. By genetic encoding of p-azido-l-phenylalanine, the protein of interest can be selectively captured using copper-free click chemistry. We use Click-MS to enrich proteins that function in different cellular compartments, and we identify protein-protein interactions, showing the great potential of Click-MS for interaction proteomics workflows.

PMID:
27643597
DOI:
10.1021/acschembio.6b00520
[Indexed for MEDLINE]

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