Study offers detailed molecular map of interaction patterns of GPCR and different G protein subtypes

G protein-coupled receptors (GPCRs) have crucial roles to play in cell signal transduction and can be used as vital therapeutic targets for several diseases.

​When GPCRs bind with extracellular agonists, they trigger several signaling pathways by employing different G proteins (G_s, G_i, G_q, etc.) to induce a wide array of physiological functions.

The biological action of the receptor depends largely on the selective coupling of a GPCR and particular G proteins.

Insights into the mechanisms of GPCR signal transduction have been limited since the molecular details characterizing how an individual GPCR identifies different G protein subtypes remain elusive.

Two cryo-electron microscopy (cryo-EM) structures of the human glucagon receptor (GCGR) together with its relative agonist glucagon and different classes of G proteins, G_s or G_i, were determined by a research team headed by WU Beili and ZHAO Qiang at the Shanghai Institute of Materia Medica (SIMM) of the Chinese Academy of Sciences (CAS), a team led by SUN Fei at the Institute of Biophysics of CAS, and a team guided by Denise Wootten from Monash University.

The study was published on March 20^th, 2020, in the Science journal.

A comprehensive molecular map of patterns of interaction between a GPCR and different G protein subtypes is provided by these structures, for the first time, surprisingly revealing several molecular features that control G protein specificity. This significantly improves the insights into GPCR signaling mechanisms.

GCGR belongs to the class B GPCR family and is vital for glucose homeostasis by initiating the release of glucose from the liver. This makes it a prospective drug target for obesity and type 2 diabetes.

GCGR canonically applies its physiological action via G_s signaling, yet it can bind to other G proteins such as G_i and G_q, thus enabling diverse cellular responses.

The crystal structures of the full-length GCGR coupled to a negative allosteric modulator or a partial peptide agonist was identified by researchers at SIMM in 2017 and 2018. That study offered an understanding of signal recognition and modulation of class B GPCRs.

Now, the researchers have achieved further progress by resolving the complex structures of GCGR attached to two transducer proteins with contradictory biological activities. This research offers a valuable understanding of pleiotropic GPCR-G protein coupling and G protein specificity.

Most importantly, it showed that GCGR’s sixth transmembrane helix (helix VI) goes through a similar outward shift in the two G protein-bound GCGR structures, thereby creating a common binding cavity to house G_s and G_i.

This is in contrast to the theory based on the previously identified GPCR-G protein complex structures, which hypothesized that the difference in positions of helix VI is a crucial determinant in the coupling specificity of G_s and G_i.

The common G protein coupling pocket visualized in the complex structures of the GCGR-G protein is consistent with the signaling pleiotropy of GCGR and enables ultimate efficiency in the activation of different pathways.

This common pocket is used by GCGR to bind to both G proteins, but GCGR performs this action in various interaction patterns accounting for G protein specificity. The quantified interaction interface between G_s and GCGR is considerably larger than it is for G_i, which leads to the increased binding affinity of G_s to the receptor. This provides a structural foundation for the preferential binding of GCGR to G_s.

The researchers took the structures of GCGR-G_s and GCGR-G_i complexes as a basis and carried out wide-scale functional studies with the help of techniques such as G protein activation, mutagenesis, and cell signaling to analyze the roles of main residues in the receptor-G protein coupling interface during G_s and G_i activation.

The study outcomes reveal that conformational variations of residue side chains and intracellular loops in the receptor are enough to guide G protein selectivity. The interactions brought about by the helix VII/VIII junction and the second intracellular loop (ICL2) of the receptor have a critical role in G_s coupling.

By contrast, ICL1 and ICL3—the other two intracellular loops—and the receptor hydrophobic intracellular binding cavity are highly crucial for G_i recognition.

These results offer in-depth knowledge about pleiotropic coupling, GPCR activation, and G protein specificity. They also offer new opportunities for drug discovery by developing biased ligands for the selective inhibition of one particular signaling pathway, which leads to reduced side effects.