Small molecule interactome mapping by photo-affinity labeling (SIM-PAL)

Non-steroidal anti-inflammatory drugs (NSAIDs) are one of the most common pharmaceutical agents in the world. However, despite well-studied interactions with cyclooxygenase-2, little is known about the broader proteomic interactions and additional mechanisms that the NSAIDs elicit. Here we present a global binding site identification platform for the direct in cellulo characterization of NSAID binding sites. The platform uses (1) photochemical conjugation of NSAID derivatives in the whole proteome, (2) enrichment of the binding sides, for (3) targeted mass spectrometry-based assignment. Using our approach, we identified the NSAID “interactome” consisting of nearly 200 binding sites in Jurkat and K562 cells. We observed a binding site hotspot on the nucleosome where three NSAIDs (but not fragment-based small molecules) interacted. These data suggest that the NSAIDs may behave as a novel type of epigenetic mark. Our approach is potentially amenable to production of precise, whole proteome binding site maps for virtually any molecule of interest.