Chemical Proteomics

Volume 803 of the series Methods in Molecular Biology pp 129-140


Identifying Cellular Targets of Small-Molecule Probes and Drugs with Biochemical Enrichment and SILAC

  • Shao-En OngAffiliated withProteomics Platform, The Broad Institute of MIT and Harvard Email author 
  • , Xiaoyu LiAffiliated withChemical Biology platform, The Broad Institute of MIT and Harvard
  • , Monica SchenoneAffiliated withProteomics platform, The Broad Institute of MIT and Harvard
  • , Stuart L. SchreiberAffiliated withChemical Biology Platform & Chemical Biology Program, The Broad Institute of MIT and Harvard
  • , Steven A. CarrAffiliated withProteomics platform, The Broad Institute of MIT and Harvard

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Sequencing of the human genome in the last decade has not yet led to a concomitant increase in the numbers of novel drug targets. While the pharmaceutical industry has invested heavily in improving drugs for existing protein targets, it has not tended toward a similar investment in experimental approaches to identify cellular targets of drugs. It is striking that the targets of numerous widely used FDA-approved drugs remain unknown. The development of robust, unbiased methods for target identification would greatly enhance our understanding the mechanisms-of-action of small molecules. Cell-based phenotypic screens followed by unbiased target identification have the potential to identify novel combinations of small molecules and their protein targets, shed light on drug polypharmacology, and enable unbiased screening approaches to drug discovery. Classical biochemical enrichment with immobilized small molecules has been used for over four decades but has been limited by issues concerning specificity and sensitivity. The application of mass spectrometry-based quantitative proteomics in combination with these affinity reagents has proven to be especially useful in addressing these common issues in affinity purification experiments. We describe the use of SILAC in identifying proteins that bind small-molecule probes and drugs in a cellular context.

Key words

Drug-target identification Proteomics SILAC Quantitation Small molecule Affinity chromatography