Surface Plasmon Resonance Study of G Protein/Receptor Coupling in a Lipid Bilayer-Free System

2006/01/11 by Konstantin E. Komolov, Ivan I. Senin, Pavel P. Philippov +2 · 45 citations
Biochemistry, Genetics and Molecular Biology · Chemistry · Neuroscience · #Biochemistry #Biophysics #Biosensor #Chemistry #Dissociation constant #G protein #G protein-coupled receptor #Lipid Membrane Structure and Behavior #Lipid bilayer #Membrane #Nanodisc #Nanoparticle #Nanotechnology #Photoreceptor and optogenetics research #Receptor #Receptor Mechanisms and Signaling #Rhodopsin #Surface plasmon resonance #Transducin

paper · doi:10.1021/ac051629t

published in Analytical Chemistry 78(4), 1228-1234 (American Chemical Society)

openalex publication_date 2006/01/11 · openalex created_date 2025/10/10 · openalex updated_date 2026/07/26

Abstract

Surface plasmon resonance (SPR) spectroscopy is a technique to study protein-protein interactions in real time; however, application of SPR spectroscopy for investigations of membrane receptors is difficult with respect to functional and uniform immobilization of receptors on a biosensor surface. In the current study, we developed a simple, direct, biosensor-based approach to monitor the molecular interactions between G protein transducin (Gt) and rhodopsin (Rho), a prototypical G protein-coupled receptor (GPCR). Detergent-solubilized dark-adapted Rho was captured onto a biosensor surface via lectin interaction, enabling site-directed immobilization of the receptor that made its cytoplasmic surface accessible to a coupling G protein. The system resembled the natural system with respect to receptor density, binding of Gt following flash or constant light application, fast GTP-dependent dissociation of Gt from Rho, regeneration of Rho, and dependence of Gt binding on light intensity and on concentration of Gt. The apparent KD of the Gt/Rho interaction was 13.6 nM. Our results validate the use of SPR spectroscopy as a tool to study G protein activation in GPCR systems and could be extended for application to other interaction partners of GPCRs.

Citations

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• Challenges in the Development of Functional Assays of Membrane Proteins [europepmc]

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