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Site-specific azide-acetyllysine photochemistry on epigenetic readers for interactome profiling

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Abstract

Chemical modifications on DNA, RNA and histones are recognized by an array of ‘reader’ modules to regulate transcriptional programming and cell fate. However, identification of reader-specific interacting partners in a dynamic cellular environment remains a significant challenge. Herein, we report a chemoproteomic approach termed ‘interaction-based protein profiling’ (IBPP) to characterize novel interacting partners of potentially any reader protein. IBPP harnesses a photosensitive amino acid introduced into the hydrophobic pocket of a reader module to crosslink and enrich transient interacting partners that are inaccessible to traditional methods. Using bromodomain-containing protein 4 (BRD4) as a paradigm, we engineer an ‘aromatic cage’ of the bromodomain to introduce 4-azido-L-phenylalanine (pAzF) without compromising its ability to recognize acetylated lysine residues in histone proteins. We establish the binding efficiency, substrate specificity and crosslinking ability of the engineered ‘reader’ module in biochemical assays. Applying IBPP, we uncovered novel acetylated interacting partners of BRD4, such as transcription factors, expanding on its previously unappreciated role in diverse biological processes. By setting up an azide-acetyllysine photoreaction deep inside the bromodomain aromatic cage as a means to detect protein acetylation, our approach provides a potentially general platform for rapid and unbiased profiling of interacting partners of diverse epigenetic readers whose functions in eukaryotic gene regulation remain convoluted.

Graphical abstract: Site-specific azide-acetyllysine photochemistry on epigenetic readers for interactome profiling

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Publication details

The article was received on 20 Jan 2017, accepted on 13 Mar 2017 and first published on 14 Mar 2017


Article type: Edge Article
DOI: 10.1039/C7SC00284J
Citation: Chem. Sci., 2017, Advance Article
  • Open access: Creative Commons BY license
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    Site-specific azide-acetyllysine photochemistry on epigenetic readers for interactome profiling

    B. Sudhamalla, D. Dey, M. Breski, T. Nguyen and K. Islam, Chem. Sci., 2017, Advance Article , DOI: 10.1039/C7SC00284J

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