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

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• 作者：Arne H. Smits ; Annika Borrmann ; Mark Roosjen ; Jan C.M. van Hest ; Michiel Vermeulen
• 刊名：ACS Chemical Biology
• 出版年：2016
• 出版时间：December 16, 2016
• 年：2016
• 卷：11
• 期：12
• 页码：3245-3250
• 全文大小：455K
• ISSN：1554-8937

文摘

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-class="smallcaps">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.