The GPCR Network: a large-scale collaboration to determine human GPCR structure and function

Abstract

G protein-coupled receptors (GPCRs) are targeted by ∼30-40% of marketed drugs, and their key roles in normal physiology and in disease demonstrate that an understanding of their structure and function is valuable to researchers in both basic science and drug discovery. However, until recently, detailed structural information on this protein family was limited by challenges in X-ray crystallographic analysis of such membrane proteins. The GPCR Network was created in 2010 with the goal of structurally characterizing 15-25 representative human GPCRs within 5 years, based on an active outreach programme addressing an interdisciplinary community of scientists interested in GPCR structure, chemistry and biology. Here, we provide an overview of how this collaborative effort has enabled the structural determination and characterization of eight human GPCRs so far, and discuss some of the challenges that remain in gaining more detailed insights into structure-function relationships in this receptor superfamily.

Figure 1

Phylogenetic tree representation of the human GPCR superfamily constructed using sequence similarity within the seven-transmembrane region. Family members with determined structures are highlighted within the tree, and their binding pockets with the ligand, as captured in each of the distinct structures, are shown around the tree in the same orientation for ease of comparison. A_2AAR (PDB code: 3EML), β₁AR (2VT4), β₂AR (2RH1), CXCR4 (3ODU), dopamine D3 (3PBL), δ-opioid (4EJ4), histamine H₁ (3RZE), κ-opioid (4DJH), µ-opioid (4DKL), M2 muscarinic (3UON), M3 muscarinic (4DAJ), nociceptin/ophanin FQ peptide opioid (4EA3), rhodopsin (1GZM), sphingolipid S1P₁ (3V2Y) receptors.

Figure 2

Process pipeline used by the GPCR Network to determine receptor structure and develop a deeper understanding of receptor dynamics and functional behavior. The stage-gate process relies on a set of metrics to advance target constructs for further processing. The bold black lines describe the process pathway; the gray lines are feedback loops where information at one stage can be used to repeat or adjust an earlier set of experiments. For example, if we find a construct is not stable enough or does not pass one of our pre-crystallization screens, we would return (gray lines) to construct design or search for more ligands that might help further stabilize the receptor.

Figure 3

The strategy of the GPCR Network includes leveraging each experimentally determined receptor structure (e.g. human β₂AR; center) towards the understanding of GPCR family diversity and receptor selectivity for ligands (right side) and individual GPCR structure-function and conformational selectivity by ligands (left side). By creating a complete data package for each receptor that includes complementary biophysical, structural, functional, and ligand data, the receptors can be more thoroughly understood in contrast to just solving the structure with limited follow up.

Figure 4

Community-wide prediction of GPCR-ligand docking and receptor interactions. In 2008 (A_2AAR in complex with ZM241385) and 2010 (dopamine D3 in complex with eticlopride; CXCR4 chemokine in complex with IT1t and CVX15), two community wide assessments were conducted where the participants submitted prediction models of undisclosed GPCR ligand structures that were recently determined by the GPCR Network. The shaded plot background represents the distribution of the corresponding parameters for pairs of symmetry-related molecules in a subset of each PDB structure. (A) 2008 GPCR Dock with the assessment of the human A_2AAR in complex with the antagonist ZM241385 highlighting limited success in receptor-ligand interactions. (B) 2010 GPCR Dock human dopamine D3 in complex with the antagonist eticlopride showing significant improvements in receptor-ligand docking, likely due to improved experimental models for the biogenic amine receptor subfamily. (C) 2010 GPCR Dock human CXCR4 receptor in complex with the small molecule IT1t. (D) 2010 GPCR Dock human CXCR4 receptor in complex with the peptide molecule CVX15. Panels A and B shows progress in the field based on available structural templates of related receptors, while panels C and D highlight the need for additional structural coverage needed for peptide receptors.

Figure 5

Receptor diversity in class A GPCRs, as illustrated by a simplified cartoon. Comparison of experimental GPCR structures reveals a larger structural diversity in the extracellular module (colored red) than in the intracellular module (colored blue), which seems to reflect the evolutionary pressure of recognizing hundreds of endogenous ligands while transferring their signals to only dozens of interacting partners (see the text for details).