Related Experiment Video
Updated: Jun 1, 2026

Measuring G-protein-coupled Receptor Signaling via Radio-labeled GTP Binding
Published on: June 9, 2017
[³⁵S]GTPγS binding as an index of total G-protein and Gα-subtype-specific activation by GPCRs
Rajendra Mistry1, Mark R Dowling, R A John Challiss
1Department of Cell Physiology and Pharmacology, University of Leicester, Leicester, UK.
Abstract:
On activation, G-protein-coupled receptors (GPCRs) exert many of their cellular actions through -promoting guanine nucleotide exchange on Gα subunits of heterotrimeric G proteins to release Gα-GTP and free βγ-subunits. In membrane preparations, GTP can be substituted by ³⁵S-labeled guanosine- 5'-O-(3-thio)triphosphate ([³⁵S]GTPγS) and on agonist stimulation a quasi-stable [³⁵S]GTPγS-Gα -complex forms and accumulates. Separation of [³⁵S]GTPγS-Gα complexes from free [³⁵S]GTPγS allows differences between basal and agonist-stimulated rates of [³⁵S]GTPγS-Gα complex formation- to be used to obtain pharmacological information on receptor-G-protein information transfer. Further, by releasing Gα-subunits into solution following the [³⁵S]GTPγS binding step, Gα-subunit-specific antibodies can be used to investigate the Gα-protein subpopulations activated by receptors by immunoprecipitation of [³⁵S]GTPγS-Gα complexes and quantification by scintillation counting. Here, we describe a total [³⁵S]GTPγS binding assay and a modification of this method that incorporates a Gα-specific immunoprecipitation step.
More Related Videos
07:41A Kinetic Fluorescence-based Ca2+ Mobilization Assay to Identify G Protein-coupled Receptor Agonists, Antagonists, and Allosteric Modulators
Published on: February 20, 2018
09:21BRET-based G Protein Biosensors for Measuring G Protein-Coupled Receptor Activity in Live Cells
Published on: November 7, 2025
Related Concept Videos
Activation and Inactivation of G Proteins
Transducer Mechanism: G Protein–Coupled Receptors
GPCRs are also called heptahelical, 7TM, or...
G Protein-coupled Receptors
GPCRs are also called heptahelical, 7TM, or serpentine receptors, and consist of seven (H1-H7) transmembrane alpha-helices that span the bilayer to form a cylindrical core. The transmembrane helices are connected by three extracellular loops and three...
G Protein-coupled Receptors
GPCRs are also called heptahelical, 7TM, or serpentine receptors, and consist of seven (H1-H7) transmembrane alpha-helices that span the bilayer to form a cylindrical core. The transmembrane helices are connected by three extracellular loops and three...
G-protein Coupled Receptors
G-protein Coupled Receptors