title: Computational analysis of the CB1 carboxyl-terminus in the receptor-G protein complex.
abstract: Despite the important role of the carboxyl-terminus (Ct) of the activated brain cannabinoid receptor one (CB1) in the regulation of G protein signaling, a structural understanding of interactions with G proteins is lacking. This is largely due to the highly flexible nature of the CB1 Ct that dynamically adapts its conformation to the presence of G proteins. In the present study, we explored how the CB1 Ct can interact with the G protein by building on our prior modeling of the CB1-Gi complex (Shim, Ahn, and Kendall, The Journal of Biological Chemistry 2013;288:32449-32465) to incorporate a complete CB1 Ct (Glu416(Ct)-Leu472(Ct)). Based on the structural constraints from NMR studies, we employed ROSETTA to predict tertiary folds, ZDOCK to predict docking orientation, and molecular dynamics (MD) simulations to obtain two distinct plausible models of CB1 Ct in the CB1-Gi complex. The resulting models were consistent with the NMR-determined helical structure (H9) in the middle region of the CB1 Ct. The CB1 Ct directly interacted with both Gα and Gβ and stabilized the receptor at the Gi interface. The results of site-directed mutagenesis studies of Glu416(Ct), Asp423(Ct), Asp428(Ct), and Arg444(Ct) of CB1 Ct suggested that the CB1 Ct can influence receptor-G protein coupling by stabilizing the receptor at the Gi interface. This research provided, for the first time, models of the CB1 Ct in contact with the G protein.
authors.affiliation: Department of Chemistry, University of North Carolina, Chapel Hill, North Carolina, 27514.
authors.forename: Joong-Youn
authors.initials: JY
authors.lastname: Shim
authors.affiliation: Department of Pharmaceutical Sciences, University of Connecticut, Storrs, Connecticut, 06269-3092.
authors.forename: Leepakshi
authors.initials: L
authors.lastname: Khurana
authors.affiliation: Department of Pharmaceutical Sciences, University of Connecticut, Storrs, Connecticut, 06269-3092.
authors.forename: Debra A
authors.initials: DA
authors.lastname: Kendall
mesh_terms: D000595:Amino Acid Sequence; D000700:Analgesics / Q000737:chemistry; D001665:Binding Sites; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D003511:Cyclohexanols / Q000737:chemistry; D019204:GTP-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000737:chemistry; D057809:HEK293 Cells; D006801:Humans; D062105:Molecular Docking Simulation; D056004:Molecular Dynamics Simulation; D016297:Mutagenesis, Site-Directed; D019906:Nuclear Magnetic Resonance, Biomolecular; D011485:Protein Binding; D000072417:Protein Domains; D017433:Protein Structure, Secondary; D021122:Protein Subunits / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D043884:Receptor, Cannabinoid, CB1 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D000700:Analgesics; D003511:Cyclohexanols; D021122:Protein Subunits; D043884:Receptor, Cannabinoid, CB1; D011994:Recombinant Proteins; D016244:Guanosine 5'-O-(3-Thiotriphosphate); C054649:3-(2-hydroxy-4-(1,1-dimethylheptyl)phenyl)-4-(3-hydroxypropyl)cyclohexanol; D019204:GTP-Binding Proteins
keywords: G protein coupling; G protein-coupled receptor; cannabinoid receptor; carboxyl-terminus; molecular dynamics; protein structure; site-directed mutagenesis