title: Distinct CCK-2 receptor conformations associated with β-arrestin-2 recruitment or phospholipase-C activation revealed by a biased antagonist.
abstract: Seven-transmembrane receptors (7TMRs), also termed G protein-coupled receptors (GPCRs), form the largest class of cell surface membrane receptors, involving several hundred members in the human genome. Nearly 30% of marketed pharmacological agents target 7TMRs. 7TMRs adopt multiple conformations upon agonist binding. Biased agonists, in contrast to non-biased agonists, are believed to stabilize conformations preferentially activating either G-protein- or β-arrestin-dependent signaling pathways. However, proof that cognate conformations of receptors display structural differences within their binding site where biased agonism initiates, are still lacking. Here, we show that a non-biased agonist, cholecystokinin (CCK) induces conformational states of the CCK2R activating Gq-protein-dependent pathway (CCK2R(G)) or recruiting β-arrestin2 (CCK2R(β)) that are pharmacologically and structurally distinct. Two structurally unrelated antagonists competitively inhibited both pathways. A third ligand (GV150013X) acted as a high affinity competitive antagonist on CCK2R(G) but was nearly inefficient as inhibitor of CCK2R(β). Several structural elements on both GV150013X and in CCK2R binding cavity, which hinder binding of GV150013X only to the CCK2R(β) were identified. At last, proximity between two conserved amino acids from transmembrane helices 3 and 7 interacting through sulfur-aromatic interaction was shown to be crucial for selective stabilization of the CCK2R(β) state. These data establish structural evidence for distinct conformations of a 7TMR associated with β-arrestin-2 recruitment or G-protein coupling and validate relevance of the design of biased ligands able to selectively target each functional conformation of 7TMRs.
authors: EA 4552, Université de Toulouse 3, 31432 Toulouse, France.
authors: Rémi
authors: R
authors: Magnan
authors: Chantal
authors: C
authors: Escrieut
authors: Véronique
authors: V
authors: Gigoux
authors: Kavita
authors: K
authors: De
authors: Pascal
authors: P
authors: Clerc
authors: Fan
authors: F
authors: Niu
authors: Joelle
authors: J
authors: Azema
authors: Bernard
authors: B
authors: Masri
authors: Arnau
authors: A
authors: Cordomi
authors: Michel
authors: M
authors: Baltas
authors: Irina G
authors: IG
authors: Tikhonova
authors: Daniel
authors: D
authors: Fourmy
mesh_terms: D000218:Adamantane / Q000031:analogs & derivatives / Q000494:pharmacology; D019390:Arrestins / Q000737:chemistry*; D001665:Binding Sites; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D018613:Microscopy, Confocal; D008958:Models, Molecular; D015394:Molecular Structure; D009154:Mutation; D010671:Phenylurea Compounds / Q000494:pharmacology; D011487:Protein Conformation; D043887:Receptor, Cholecystokinin B / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics; D015398:Signal Transduction; D010738:Type C Phospholipases / Q000737:chemistry / Q000378:metabolism*; D015854:Up-Regulation; D000071560:beta-Arrestin 2; D000071557:beta-Arrestins
chemical_list: C000605254:ARRB2 protein, human; D019390:Arrestins; C111554:GV 150013X; D010671:Phenylurea Compounds; D043887:Receptor, Cholecystokinin B; D000071560:beta-Arrestin 2; D000071557:beta-Arrestins; D010738:Type C Phospholipases; D000218:Adamantane