Daisuke Ogasawara, David Konrad, Zher Yin Tan, Kimberly Carey, Jessica Luo, Sang Joon Won, Haoxin Li, Trever Carter, Kristen DeMeester, Evert Njomen, Stuart Schreiber, Ramnik Xavier, Bruno Melillo, Benjamin Cravatt
Cell Chemical Biology, 2024
https://doi.org/10.1016/j.chembiol.2024.10.005
bioRxiv 2024.02.27.582206;
doi: https://doi.org/10.1101/2024.02.27.582206
Chemical proteomics has emerged as a versatile approach to discover small-molecule ligands for proteins in native biological systems. Nonetheless, only a limited diversity of chemistry has so far been explored in chemical proteomic studies. In this study, we describe a set of DOS-inspired “photo-stereoprobes” that possess distinct physicochemical properties (e.g., greater three-dimensionality) compared to fragments. We identified >200 proteins showing stereoselective interactions with photo-stereoprobes in human cancer cells and confirmed many of these binding events with recombinant proteins. Interestingly, interactions that were not recapitulated in recombinant systems were enriched for proteins that are parts of large complexes, and additional stereoselective liganding events were identified exclusively in cells, but not cell lysates. These results highlight the value of chemical proteomic experiments that can map small-molecule interactions with endogenously expressed proteins in living systems. More broadly, the stereoprobe-liganded proteins were from diverse structural and functional classes, including adaptor/scaffolding and transcriptional regulatory proteins that have been historically challenging to target with small molecules. We further demonstrated that photo-stereoprobe-protein interactions can be converted into NanoBRET assays compatible with HTS to enable access to larger and more diverse small-molecule libraries. Finally, we demonstrated the utility of photo-stereoprobes in phenotypic screening by discovering compounds that stereoselectively modulate autophagy through engaging the mitochondrial protease CLPP. Taken together, our findings support the versatility of photo-stereoprobes as tools to expand the ligandability of the human proteome.
