In situ chemoproteomic profiling reveals itaconate inhibits de novo purine biosynthesis in pathogens

Itaconate serves as an immune-specific metabolite that regulates gene transcription and metabolism in both host and pathogens. S-itaconation is a post-translational modification that regulates immune response; however, its antimicrobial mechanism under the physiological condition remains unclear. Here, we apply a bioorthogonal itaconate probe to perform global profiling of S-itaconation in living pathogens, including S. Typhimurium, S. aureus, and P. aeruginosa. Some functional enzymes are covalently modified by itaconate, including those involved in the de novo purine biosynthesis pathway. Further biochemical studies demonstrate that itaconate suppresses this specific pathway to limit Salmonella growth by inhibiting the initiator purF to lower de novo purine biosynthesis and simultaneously targeting the guaABC cluster to block the salvage route. Our chemoproteomic study provides a global portrait of S-itaconation in multiple pathogens and offers a valuable resource for finding susceptible targets to combat drug-resistant pathogens in the future. [Display omitted] •S-itaconation is quantitatively profiled in three living pathogens by a bioorthogonal probe•Itaconate covalently modifies functional cysteines of distinct enzymes in three bacteria•Itaconate inhibits key enzymes of de novo purine biosynthesis to suppress Salmonella growth Itaconate has multifaced functions in the interactions between host and pathogens. Liu et al. employ a bioorthogonal probe and quantitative chemoproteomics to globally profile itaconate-modified cysteines in three living pathogens, showing that itaconate covalently inhibits key enzymes of de novo purine biosynthesis in pathogenic Salmonella to suppress bacterial growth.