title: Allosteric coupling from G protein to the agonist-binding pocket in GPCRs.
abstract: G-protein-coupled receptors (GPCRs) remain the primary conduit by which cells detect environmental stimuli and communicate with each other. Upon activation by extracellular agonists, these seven-transmembrane-domain-containing receptors interact with heterotrimeric G proteins to regulate downstream second messenger and/or protein kinase cascades. Crystallographic evidence from a prototypic GPCR, the β2-adrenergic receptor (β2AR), in complex with its cognate G protein, Gs, has provided a model for how agonist binding promotes conformational changes that propagate through the GPCR and into the nucleotide-binding pocket of the G protein α-subunit to catalyse GDP release, the key step required for GTP binding and activation of G proteins. The structure also offers hints about how G-protein binding may, in turn, allosterically influence ligand binding. Here we provide functional evidence that G-protein coupling to the β2AR stabilizes a ‘closed’ receptor conformation characterized by restricted access to and egress from the hormone-binding site. Surprisingly, the effects of G protein on the hormone-binding site can be observed in the absence of a bound agonist, where G-protein coupling driven by basal receptor activity impedes the association of agonists, partial agonists, antagonists and inverse agonists. The ability of bound ligands to dissociate from the receptor is also hindered, providing a structural explanation for the G-protein-mediated enhancement of agonist affinity, which has been observed for many GPCR–G-protein pairs. Our data also indicate that, in contrast to agonist binding alone, coupling of a G protein in the absence of an agonist stabilizes large structural changes in a GPCR. The effects of nucleotide-free G protein on ligand-binding kinetics are shared by other members of the superfamily of GPCRs, suggesting that a common mechanism may underlie G-protein-mediated enhancement of agonist affinity.
authors.forename: Brian T
authors.initials: BT
authors.lastname: DeVree
authors.forename: Jacob P
authors.initials: JP
authors.lastname: Mahoney
authors.forename: Gisselle A
authors.initials: GA
authors.lastname: Vélez-Ruiz
authors.forename: Soren G F
authors.initials: SG
authors.lastname: Rasmussen
authors.forename: Adam J
authors.initials: AJ
authors.lastname: Kuszak
authors.forename: Elin
authors.initials: E
authors.lastname: Edwald
authors.forename: Juan-Jose
authors.initials: JJ
authors.lastname: Fung
authors.forename: Aashish
authors.initials: A
authors.lastname: Manglik
authors.forename: Matthieu
authors.initials: M
authors.lastname: Masureel
authors.forename: Yang
authors.initials: Y
authors.lastname: Du
authors.forename: Rachel A
authors.initials: RA
authors.lastname: Matt
authors.forename: Els
authors.initials: E
authors.lastname: Pardon
authors.forename: Jan
authors.initials: J
authors.lastname: Steyaert
authors.forename: Brian K
authors.initials: BK
authors.lastname: Kobilka
authors.forename: Roger K
authors.initials: RK
authors.lastname: Sunahara
mesh_terms: D058666:Adrenergic beta-2 Receptor Agonists / Q000378:metabolism; D058672:Adrenergic beta-2 Receptor Antagonists / Q000378:metabolism; D000494:Allosteric Regulation / Q000187:drug effects; D000495:Allosteric Site / Q000187:drug effects; D019205:GTP-Binding Protein alpha Subunits, Gs / Q000378:metabolism* / Q000494:pharmacology; D006147:Guanine / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D008024:Ligands; D008958:Models, Molecular; D011485:Protein Binding / Q000187:drug effects; D011487:Protein Conformation / Q000187:drug effects; D018343:Receptors, Adrenergic, beta-2 / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism*; D057127:Single-Chain Antibodies / Q000276:immunology / Q000494:pharmacology
chemical_list: D058666:Adrenergic beta-2 Receptor Agonists; D058672:Adrenergic beta-2 Receptor Antagonists; D008024:Ligands; C000609472:Nb80 nanobody; D018343:Receptors, Adrenergic, beta-2; D057127:Single-Chain Antibodies; D006147:Guanine; D019205:GTP-Binding Protein alpha Subunits, Gs