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- Title
Fluorescence ratiometric detection of ligand-induced receptor internalization using extracellular coiled-coil tag–probe labeling
- Authors
Yano, Yoshiaki; Matsuzaki, Katsumi
- Abstract
Abstract: We report a new method for the detection of ligand-induced receptor internalization by fluorescence ratiometric imaging of pH in endosomes in combination with a recently developed posttranslational labeling system based on the formation of a heterodimeric coiled-coil structure. The N-terminus of the β2-adrenergic receptor expressed on the cell surface was doubly labeled with pH-sensitive fluorescein and pH-insensitive tetramethylrhodamine. A significant increase in the tetramethylrhodamine-to-fluorescein fluorescence intensity ratio was observed after incubation with agonists in a concentration-dependent manner. This simple and accurate method of detecting the agonistic activity of receptors will be useful for high-throughput screening of drug candidates.
- Subjects
FLUORESCENCE; LIGANDS (Biochemistry); CELL receptors; ENDOCYTOSIS; ADRENERGIC receptors; FLUORESCEIN; ENDOSOMES; PH effect
- Publication
FEBS Letters, 2011, Vol 585, Issue 14, p2385
- ISSN
0014-5793
- Publication type
Article
- DOI
10.1016/j.febslet.2011.06.012