Sulfonyl-purine (SuPUR) chemoproteomics enables regioselective targeting of the human purine interactome, capturing 31,000+ ligandable tyrosine and lysine sites that can be rapidly developed into potent, proteome-wide selective inhibitors through fragment-based approaches, with demonstrated applications including non-catalytic serine hydrolase inhibition, dual catalytic/PPI disruption of multifunctional enzymes, and phenotypic discovery of ACAT2 as a metabolic dependency in cancer cells.

Purines are essential bioactive molecules that interact with a large fraction of the human proteome. Despite their importance, the scope of actionable purine-binding pockets for ligand discovery remains limited. Here, we developed a quantitative chemoproteomics platform using sulfonyl-purine (SuPUR) chemistry to produce a massive and functional map of the human purine interactome. The SuPUR platform captured 31,000+ targetable tyrosine and lysine sites, representing the most comprehensive beyond cysteine chemoproteomics database for enabling protein ligand discovery. SuPUR ligands that bind through a regioselective fashion serve as enabling starting points for developing potent (nanomolar) and proteome-wide-selective modulators of enzymatic and protein-protein interaction function. Phenotypic screening identified a site-specific (Y237) and regioselective SuPUR ligand of ACAT2 to reveal an unexpected metabolic dependency in cancer cells. A crystal structure of SuPUR ligand-bound ACAT2 revealed the purine group binds deep in the CoA pocket forming key interactions with catalytic residues via a water bridge to guide future structure-based ligand design.