Abstract
Understanding how small molecules bind to specific protein complexes in living cells is critical to understanding their mechanism-of-action. Unbiased chemical biology strategies for direct readout of protein interactome remodelling by small molecules would provide advantages over target-focused approaches, including the ability to detect previously unknown ligand targets and complexes. However, there are few current methods for unbiased profiling of small molecule interactomes. To address this, we envisioned a technology that would combine the sensitivity and live-cell compatibility of proximity labelling coupled to mass spectrometry, with the specificity and unbiased nature of chemoproteomics. In this manuscript, we describe the BioTAC system, a small-molecule guided proximity labelling platform that can rapidly identify both direct and complexed small molecule binding proteins. We benchmark the system against µMap, photoaffinity labelling, affinity purification coupled to mass spectrometry and proximity labelling coupled to mass spectrometry datasets. We also apply the BioTAC system to provide interactome maps of Trametinib and analogues. The BioTAC system overcomes a limitation of current approaches and supports identification of both inhibitor bound and molecular glue bound complexes.