A Modular Fluorescent Sensor Motif Used to Detect Opioids, Protein–Protein Interactions, and Protease Activity

Modular fluorescent sensor motifs are needed to design fluorescent sensors for detecting various cellular processes and functional molecules. Here, we took advantage of the versatility of a new sensor motif to design a series of sensors called SPOTon. SPOTon sensors integrate the signal from either opioids, protein–protein interactions, or protease activities to generate persistent green fluorescence. We demonstrate that SPOTon can be engineered with temporal gating to allow detection of these cellular events during a user-defined time window, providing temporal information about cellular processes and functional molecule release. These SPOTon sensors all show a high signal-to-noise ratio, up to 38 for chemical gated opioid detection, 147 for chemical gated protein–protein interaction detection, and 85 for protease activity detection.