Conformational changes in the G protein Gs induced by the β2 adrenergic receptor

Abstract

G-protein-coupled receptors (GPCRs) mediate the majority of a cell's responses to hormones and neurotransmitters, and to the senses of sight, olfaction and taste. This makes GPCRs potentially the most important group of drug targets in the human body. GPCRs are deeply embedded in the cell membrane, crossing it seven times, so structure determination for these complexes is particularly challenging — as recounted in a recent News Feature (see http://go.nature.com/ftqnx4 ). The eagerly-awaited X-ray crystal structure of a GPCR transmembrane signalling complex has now been determined by Brian Kobilka's group. The structure presented is of an agonist-occupied monomer of the β2 adrenergic receptor in complex with Gs, the stimulatory G protein for adenylyl cyclase. An accompanying paper reports the use of peptide amide hydrogen-deuterium exchange mass spectrometry to probe the protein dynamics of this signalling complex. G protein-coupled receptors represent the largest family of membrane receptors1 that instigate signalling through nucleotide exchange on heterotrimeric G proteins. Nucleotide exchange, or more precisely, GDP dissociation from the G protein α-subunit, is the key step towards G protein activation and initiation of downstream signalling cascades. Despite a wealth of biochemical and biophysical studies on inactive and active conformations of several heterotrimeric G proteins, the molecular underpinnings of G protein activation remain elusive. To characterize this mechanism, we applied peptide amide hydrogen–deuterium exchange mass spectrometry to probe changes in the structure of the heterotrimeric bovine G protein, Gs (the stimulatory G protein for adenylyl cyclase) on formation of a complex with agonist-bound human β2 adrenergic receptor (β2AR). Here we report structural links between the receptor-binding surface and the nucleotide-binding pocket of Gs that undergo higher levels of hydrogen–deuterium exchange than would be predicted from the crystal structure of the β2AR–Gs complex. Together with X-ray crystallographic and electron microscopic data of the β2AR–Gs complex (from refs 2, 3), we provide a rationale for a mechanism of nucleotide exchange, whereby the receptor perturbs the structure of the amino-terminal region of the α-subunit of Gs and consequently alters the ‘P-loop’ that binds the β-phosphate in GDP. As with the Ras family of small-molecular-weight G proteins, P-loop stabilization and β-phosphate coordination are key determinants of GDP (and GTP) binding affinity.