title: Structural flexibility of the G alpha s alpha-helical domain in the beta2-adrenoceptor Gs complex.
abstract: The active-state complex between an agonist-bound receptor and a guanine nucleotide-free G protein represents the fundamental signaling assembly for the majority of hormone and neurotransmitter signaling. We applied single-particle electron microscopy (EM) analysis to examine the architecture of agonist-occupied β(2)-adrenoceptor (β(2)AR) in complex with the heterotrimeric G protein Gs (Gαsβγ). EM 2D averages and 3D reconstructions of the detergent-solubilized complex reveal an overall architecture that is in very good agreement with the crystal structure of the active-state ternary complex. Strikingly however, the α-helical domain of Gαs appears highly flexible in the absence of nucleotide. In contrast, the presence of the pyrophosphate mimic foscarnet (phosphonoformate), and also the presence of GDP, favor the stabilization of the α-helical domain on the Ras-like domain of Gαs. Molecular modeling of the α-helical domain in the 3D EM maps suggests that in its stabilized form it assumes a conformation reminiscent to the one observed in the crystal structure of Gαs-GTPγS. These data argue that the α-helical domain undergoes a nucleotide-dependent transition from a flexible to a conformationally stabilized state.
authors.affiliation: Life Sciences Institute and Department of Biological Chemistry, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
authors.forename: Gerwin H
authors.initials: GH
authors.lastname: Westfield
authors.forename: Søren G F
authors.initials: SG
authors.lastname: Rasmussen
authors.forename: Min
authors.initials: M
authors.lastname: Su
authors.forename: Somnath
authors.initials: S
authors.lastname: Dutta
authors.forename: Brian T
authors.initials: BT
authors.lastname: DeVree
authors.forename: Ka Young
authors.initials: KY
authors.lastname: Chung
authors.forename: Diane
authors.initials: D
authors.lastname: Calinski
authors.forename: Gisselle
authors.initials: G
authors.lastname: Velez-Ruiz
authors.forename: Austin N
authors.initials: AN
authors.lastname: Oleskie
authors.forename: Els
authors.initials: E
authors.lastname: Pardon
authors.forename: Pil Seok
authors.initials: PS
authors.lastname: Chae
authors.forename: Tong
authors.initials: T
authors.lastname: Liu
authors.forename: Sheng
authors.initials: S
authors.lastname: Li
authors.forename: Virgil L
authors.initials: VL
authors.lastname: Woods
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
authors.forename: Georgios
authors.initials: G
authors.lastname: Skiniotis
mesh_terms: D000818:Animals; D003460:Crystallization; D018360:Crystallography, X-Ray; D019205:GTP-Binding Protein alpha Subunits, Gs / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000737:chemistry / Q000378:metabolism; D006153:Guanosine Diphosphate / Q000737:chemistry / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000737:chemistry / Q000378:metabolism; D008854:Microscopy, Electron; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D018343:Receptors, Adrenergic, beta-2 / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure
chemical_list: D018343:Receptors, Adrenergic, beta-2; D006153:Guanosine Diphosphate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate; D019205:GTP-Binding Protein alpha Subunits, Gs