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Measuring G-protein-coupled Receptor Signaling via Radio-labeled GTP Binding
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1The Solomon H. Snyder Department of Neuroscience,Johns Hopkins University School of Medicine, 2Department of Neurology and neurosurgery,Johns Hopkins University School of Medicine, 3Departments of Pharmacology and Molecular Sciences,Johns Hopkins University School of Medicine, 4Department of Psychiatry and Behavioral Sciences,Johns Hopkins University School of Medicine
Chapters
- 00:05Title
- 00:37Expression of Recombinant HA-MOR1, Cell Fractionation, and Membrane Collection
- 03:55[35S]GTPγS Binding
- 06:05Membrane Filtration
- 06:59Liquid Scintillation Counting
- 07:57Results: Characterization of G-protein-coupled Receptors by Monitoring Radio-labled GTP Binding
- 09:01Conclusion
Guanosine triphosphate (GTP) binding is one of the earliest events in G-Protein-Coupled Receptor (GPCR) activation. This protocol describes how to pharmacologically characterize specific GPCR-ligand interactions by monitoring the binding of the radio-labeled GTP analog, [35S]guanosine-5'-O-(3-thio)triphosphate ([35S]GTPγS), in response to a ligand of interest.
