Structural flexibility of the G alpha s alpha-helical domain in the beta2-adrenoceptor Gs complex

Proc Natl Acad Sci U S A. 2011 Sep 20;108(38):16086-91. doi: 10.1073/pnas.1113645108. Epub 2011 Sep 13.

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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Crystallization
  • Crystallography, X-Ray*
  • GTP-Binding Protein alpha Subunits, Gs / chemistry*
  • GTP-Binding Protein alpha Subunits, Gs / metabolism
  • GTP-Binding Protein alpha Subunits, Gs / ultrastructure
  • Guanosine 5'-O-(3-Thiotriphosphate) / chemistry
  • Guanosine 5'-O-(3-Thiotriphosphate) / metabolism
  • Guanosine Diphosphate / chemistry
  • Guanosine Diphosphate / metabolism
  • Guanosine Triphosphate / chemistry
  • Guanosine Triphosphate / metabolism
  • Microscopy, Electron
  • Models, Molecular
  • Protein Binding
  • Protein Conformation
  • Protein Structure, Secondary*
  • Protein Structure, Tertiary*
  • Receptors, Adrenergic, beta-2 / chemistry*
  • Receptors, Adrenergic, beta-2 / metabolism
  • Receptors, Adrenergic, beta-2 / ultrastructure

Substances

  • Receptors, Adrenergic, beta-2
  • Guanosine Diphosphate
  • Guanosine 5'-O-(3-Thiotriphosphate)
  • Guanosine Triphosphate
  • GTP-Binding Protein alpha Subunits, Gs