Bioorthogonal photocatalytic decaging-enabled spatiotemporal proteomics

Spatiotemporally resolved dissection of subcellular proteome is crucial to our understanding of cellular functions in health and disease. Although enzyme-based proximity labeling strategies have emerged as powerful methods to portray the compartmentalized proteome of diverse organelles, current approaches still suffer from limitations such as the genetic operation that is not compatible with hard-to-transfect cells as well as overexpression of the fusion protein that may cause altered intracellular localization. We herein report a non-genetic strategy termed bioorthogonal and photocatalytic decaging-mediated proximity labeling (CAT-Prox) for spatiotemporally resolved proteome profiling in living cells. Our systematic survey of the organometallic photocatalysts has led to the identification of Ir(ppy)2bpy as a bioorthogonal and mitochondria-targeting complex that allowed photo-controlled, rapid rescue of azidobenzyl-caged quinone methide as a highly reactive Michael acceptor for proximity-based protein labelings. By coupling with quantitative mass spectrometry, CAT-Prox revealed the dynamic mitochondria proteome of cancer cells and macrophage cells under normal and stressed conditions. Furthermore, by targeting the photocatalyst to cell membrane receptors, CAT-Prox allowed microdomain proteome profiling on live cell surface. Together, CAT-Prox integrated the advantages of both enzymatic and chemical-based proximity labeling approaches as a general spatiotemporal proteomics platform for diverse subcellular spaces and cell types.