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Updated: Jun 25, 2026

Identification of Small Molecule-binding Proteins in a Native Cellular Environment by Live-cell Photoaffinity Labeling
Published on: September 20, 2016
Identifying the proteins to which small-molecule probes and drugs bind in cells
Shao-En Ong1, Monica Schenone, Adam A Margolin
1Proteomics Platform, The Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA. song@broad.mit.edu
Abstract:
Most small-molecule probes and drugs alter cell circuitry by interacting with 1 or more proteins. A complete understanding of the interacting proteins and their associated protein complexes, whether the compounds are discovered by cell-based phenotypic or target-based screens, is extremely rare. Such a capability is expected to be highly illuminating--providing strong clues to the mechanisms used by small-molecules to achieve their recognized actions and suggesting potential unrecognized actions. We describe a powerful method combining quantitative proteomics (SILAC) with affinity enrichment to provide unbiased, robust and comprehensive identification of the proteins that bind to small-molecule probes and drugs. The method is scalable and general, requiring little optimization across different compound classes, and has already had a transformative effect on our studies of small-molecule probes. Here, we describe in full detail the application of the method to identify targets of kinase inhibitors and immunophilin binders.
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