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Significance Copper is a required metal nutrient for life, yet its altered homeostasis is associated with many diseases. Thus, to develop new methods to help decipher copper biology, we present an activity-based ratiometric fluorescence resonance energy transfer (FRET) probe that exploits a biomimetic, copper(I)-dependent cleavage reaction to enable imaging of loosely bound, labile copper pools in cells with metal and oxidation state selectivity and a self-calibrating ratiometric response. Application of this technology to cellular models of cancer reveals that oncogene-driven changes in the metabolism of glutathione, a major cellular redox buffer, leads to a labile copper(I) deficiency. This work establishes the relevance of copper dysregulation to cancer metabolism and presages further opportunities for activity-based sensing in studies of metal biology.

Activity-based ratiometric FRET probe reveals oncogene-driven changes in labile copper pools induced by altered glutathione metabolism