Unique agonist-bound cannabinoid CB 1 receptor conformations indicate agonist specificity in signaling

• Published in: European journal of pharmacology Vol. 581; no. 1; pp. 19 - 29
• Main Authors: Georgieva, Teodora, Devanathan, Savitha, Stropova, Dagmar, Park, Chad K., Salamon, Zdzislaw, Tollin, Gordon, Hruby, Victor J., Roeske, William R., Yamamura, Henry I., Varga, Eva
• Format: Journal Article
• Language: English
• Published: Elsevier B.V 2008
• Subjects: Functional selectivity; G proteins; PWR spectroscopy; Trafficking; Trafficking; Functional selectivity; G proteins; PWR spectroscopy
• ISSN: 0014-2999; 1879-0712
• DOI: 10.1016/j.ejphar.2007.11.053

Cannabinoid drugs differ in their rank order of potency to produce analgesia versus other central nervous system effects. We propose that these differences are due to unique agonist-bound cannabinoid CB 1 receptor conformations that exhibit different affinities for individual subsets of intracellular signal transduction pathways. In order to test this hypothesis, we have used plasmon-waveguide resonance (PWR) spectroscopy, a sensitive method that can provide direct information about ligand–protein and protein–protein interactions, and can detect conformational changes in lipid-embedded proteins. A recombinant epitope-tagged human cannabinoid CB 1 receptor was expressed in insect Sf9 cells, solubilized and purified using two-step affinity chromatography. The purified receptor was incorporated into a lipid bilayer on the surface of the PWR resonator. PWR spectroscopy demonstrated that cannabinoid agonists exhibit high affinity ( K D = 0.2 ± 0.03 nM and 2 ± 0.4 nM for CP 55,940 and WIN 55,212-2, respectively) for the purified epitope tagged hCB 1 receptor. Interestingly however, these structurally different cannabinoid agonists shifted the PWR spectra in opposite directions, indicating that CP 55,940 and WIN 55,212-2 binding leads to different hCB 1 receptor conformations. Furthermore, PWR experiments also indicated that these CP 55,940-and WIN 55,212-bound hCB 1 receptor conformations exhibit slightly different affinities to an inhibitory G protein heterotrimer, G i1 ( K D = 27 ± 8 nM and K D = 10.7 ± 4.7 nM, respectively), whereas they strikingly differ in their ability to activate this G protein type.