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Updated: Jun 7, 2026
10:13
Measuring G-protein-coupled Receptor Signaling via Radio-labeled GTP Binding
Published on: June 9, 2017
Biacore analysis with stabilized G-protein-coupled receptors
Rebecca L Rich1, James Errey, Fiona Marshall
1Center for Biomolecular Interaction Analysis, University of Utah School of Medicine, Salt Lake City, 84132, USA.
Analytical Biochemistry
|October 26, 2010
Summary
Researchers developed a new biosensor method using stabilized G-protein-coupled receptors to accurately measure drug interactions. This technique enhances the characterization of small-molecule antagonists for β₁ adrenergic and A₂(A) adenosine receptors.
Area of Science:
- Biochemistry and Molecular Pharmacology
- Biosensor Technology
- G-Protein-Coupled Receptor (GPCR) Research
Background:
- G-protein-coupled receptors (GPCRs) are crucial drug targets, but their characterization is challenging.
- Existing methods for analyzing receptor-ligand interactions often lack stability and high capacity.
- Stabilized receptor forms are essential for reliable binding studies.
Purpose of the Study:
- To apply Biacore technology with stabilized GPCRs for precise activity monitoring.
- To characterize binding constants of small-molecule antagonists across a wide affinity range.
- To develop and validate improved methods for receptor immobilization and regeneration on biosensor chips.
Main Methods:
- Utilized stabilized β₁ adrenergic and A₂(A) adenosine G-protein-coupled receptors.
- Employed Biacore surface plasmon resonance (SPR) technology for real-time binding analysis.
- Developed an improved method for tethering His-tagged receptors on NTA-functionalized carboxymethylated dextran chips.
- Implemented a novel approach for regenerating receptor binding sites post-analysis.
Main Results:
- Successfully monitored receptor activity and characterized binding constants for small-molecule antagonists.
- Demonstrated a >20,000-fold range in antagonist affinity.
- Achieved stable, high-capacity, and high-activity receptor surfaces using the improved NTA-chip immobilization method.
- Validated a novel receptor binding site regeneration technique.
Conclusions:
- The combination of stabilized GPCRs and biosensor technology provides a robust platform for drug discovery.
- The improved immobilization and regeneration methods enhance the efficiency and reliability of receptor characterization.
- This approach is expected to become a standard method for analyzing GPCR-ligand interactions.
Related Concept Videos
01:21G-protein Coupled Receptors
G-protein coupled receptors are ligand binding receptors that indirectly affect changes in the cell. The actual receptor is a single polypeptide that transverses the cell membrane seven times creating intracellular and extracellular loops. The extracellular loops create a ligand specific pocket which binds to neurotransmitters or hormones. The intracellular loops holds onto the G-protein.
01:21G-protein Coupled Receptors
G-protein coupled receptors are ligand binding receptors that indirectly affect changes in the cell. The actual receptor is a single polypeptide that transverses the cell membrane seven times creating intracellular and extracellular loops. The extracellular loops create a ligand specific pocket which binds to neurotransmitters or hormones. The intracellular loops holds onto the G-protein.
01:22Activation and Inactivation of G Proteins
Heterotrimeric G proteins are guanine nucleotide-binding proteins. As the name suggests, heterotrimeric G proteins are composed of three subunits: alpha, beta, and gamma. They remain GDP-bound or GTP-bound inside the cells and switch between inactive/active states. The Gα subunit possesses the nucleotide-binding pocket that binds guanine nucleotides and switches between GDP or GTP-bound states. In contrast, the Gꞵ and Gγ subunits are always bound together with high affinity and are together...
01:15G Protein-coupled Receptors
G Protein-Coupled Receptors or GPCRs are membrane-bound receptors that transiently associate with heterotrimeric G proteins and induce an appropriate response to sensory stimuli such as light, odors, hormones, cytokines, or neurotransmitters.
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...
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01:15G Protein-coupled Receptors
G Protein-Coupled Receptors or GPCRs are membrane-bound receptors that transiently associate with heterotrimeric G proteins and induce an appropriate response to sensory stimuli such as light, odors, hormones, cytokines, or neurotransmitters.
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...
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G Protein–Coupled Receptors (GPCRs) are membrane-bound receptors that transiently associate with heterotrimeric G proteins and induce an appropriate response to various stimuli. GPCRs regulate critical physiological pathways and are excellent drug targets for treating diseases such as diabetes, cancer, obesity, depression, or Alzheimer's. Nearly 35% of approved drugs implement their therapeutic effects by selectively interacting with specific GPCRs.
GPCRs are also called heptahelical, 7TM, or...
GPCRs are also called heptahelical, 7TM, or...