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• W3091890131 abstract "Rhodopsin is a canonical class A photosensitive G protein-coupled receptor (GPCR), yet relatively few pharmaceutical agents targeting this visual receptor have been identified, in part due to the unique characteristics of its light-sensitive, covalently bound retinal ligands. Rhodopsin becomes activated when light isomerizes 11-cis-retinal into an agonist, all-trans-retinal (ATR), which enables the receptor to activate its G protein. We have previously demonstrated that, despite being covalently bound, ATR can display properties of equilibrium binding, yet how this is accomplished is unknown. Here, we describe a new approach for both identifying compounds that can activate and attenuate rhodopsin and testing the hypothesis that opsin binds retinal in equilibrium. Our method uses opsin-based fluorescent sensors, which directly report the formation of active receptor conformations by detecting the binding of G protein or arrestin fragments that have been fused onto the receptor's C terminus. We show that these biosensors can be used to monitor equilibrium binding of the agonist, ATR, as well as the noncovalent binding of β-ionone, an antagonist for G protein activation. Finally, we use these novel biosensors to observe ATR release from an activated, unlabeled receptor and its subsequent transfer to the sensor in real time. Taken together, these data support the retinal equilibrium binding hypothesis. The approach we describe should prove directly translatable to other GPCRs, providing a new tool for ligand discovery and mutant characterization." @default.
• W3091890131 created "2020-10-15" @default.
• W3091890131 creator A5003487863 @default.
• W3091890131 creator A5043616114 @default.
• W3091890131 creator A5051792964 @default.
• W3091890131 date "2020-12-01" @default.
• W3091890131 modified "2023-09-28" @default.
• W3091890131 title "Novel fluorescent GPCR biosensor detects retinal equilibrium binding to opsin and active G protein and arrestin signaling conformations" @default.
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• W3091890131 doi "https://doi.org/10.1074/jbc.ra120.014631" @default.
• W3091890131 hasPubMedCentralId "https://www.ncbi.nlm.nih.gov/pmc/articles/7762930" @default.
• W3091890131 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/33453993" @default.
• W3091890131 hasPublicationYear "2020" @default.
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