Switchable DNA Catalysts for Proximity Labeling at Sites of Protein-Protein Interactions

Proximitylabeling (PL) has emerged as a powerful approach to elucidateproteomes within a defined radius around a protein of interest (POI).In PL, a catalyst is attached to the POI and tags nearby endogenousproteins, which are then isolated by affinity purification and identifiedby mass spectrometry. Although existing PL methods have yielded numerousbiological insights, proteomes with greater spatial resolution couldbe obtained if PL catalysts could be activated at more specific subcellularlocations, such as sites where both the POI and a chemical stimulusare present or sites of protein-protein interactions (PPIs).Here, we report DNA-based switchable PL catalysts that are attachedto a POI and become activated only when a secondary molecular triggeris present. The DNA catalysts consist of a photocatalyst and a spectralquencher tethered to a DNA oligomer. They are catalytically inactiveby default but undergo a conformational change in response to a specificmolecular trigger, thus activating PL. We designed a system in whichthe DNA catalyst becomes activated on living mammalian cells specificallyat sites of Her2-Her3 heterodimers and c-Met homodimers, PPIsknown to increase the invasion and growth of certain cancers. Whilethis study employs a Ru(bpy)(3)-type complex for taggingproteins with biotin phenol, the switchable DNA catalyst design iscompatible with diverse synthetic PL photocatalysts. Furthermore,the switchable DNA PL catalysts can be constructed from conformation-switchingDNA aptamers that respond to small molecules, ions, and proteins,opening future opportunities for PL in highly specific subcellularlocations.