A common ligand-binding site in G-protein coupled receptors

Abstract Antibodies raised against an intracellular and extracellular domain of the GH secretagogue receptor (GHS-R) confirmed that its topological orientation in the lipid bilayer is as predicted for G protein-coupled receptors with seven transmembrane domains. A strategy for mapping the agonist-binding site of the human GHS-R was conceived based on our understanding of ligand binding in biogenic amine and peptide hormone G protein-coupled receptors. Using site-directed mutagenesis and molecular modeling, we classified GHS peptide and nonpeptide agonist binding in the context of its receptor environment. All peptide and nonpeptide ligand classes shared a common binding domain in transmembrane (TM) region 3 of the GHS-R. This finding was based on TM-3 mutation E124Q, which eliminated the counter-ion to the shared basic N+ group of all GHSs and resulted in a nonfunctional receptor. Restoration of function for the E124Q mutant was achieved by a complementary change in the MK-0677 ligand through modification of its amine side-chain to the corresponding alcohol. Contacts in other TM domains [TM-2 (D99N), TM-5 (M213K, S117A), TM-6 (H280F), and extracellular loop 1 (C116A)] of the receptor revealed specificity for the different peptide, benzolactam, and spiroindolane GHSs. GHS-R agonism, therefore, does not require identical disposition of all agonist classes at the ligand-binding site. Our results support the hypothesis that the ligand-binding pocket in the GHS-R is spatially disposed similarly to the well characterized catechol-binding site in theβ 2-adrenergic receptor.

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A structural basis for how ligand binding site changes can allosterically regulate GPCR signaling and engender functional selectivity

Science Signaling ◽

10.1126/scisignal.aaw5885 ◽

2020 ◽

Vol 13 (617) ◽

pp. eaaw5885 ◽

Cited By ~ 5

Author(s):

Marta Sanchez-Soto ◽

Ravi Kumar Verma ◽

Blair K. A. Willette ◽

Elizabeth C. Gonye ◽

Annah M. Moore ◽

...

Keyword(s):

Ligand Binding ◽

Binding Site ◽

G Protein ◽

Structural Basis ◽

Protein Signaling ◽

Ligand Binding Site ◽

Intracellular Loop ◽

Gpcr Signaling ◽

Biased Signaling ◽

Dynamics Simulations

Signaling bias is the propensity for some agonists to preferentially stimulate G protein–coupled receptor (GPCR) signaling through one intracellular pathway versus another. We previously identified a G protein–biased agonist of the D2 dopamine receptor (D2R) that results in impaired β-arrestin recruitment. This signaling bias was predicted to arise from unique interactions of the ligand with a hydrophobic pocket at the interface of the second extracellular loop and fifth transmembrane segment of the D2R. Here, we showed that residue Phe189 within this pocket (position 5.38 using Ballesteros-Weinstein numbering) functions as a microswitch for regulating receptor interactions with β-arrestin. This residue is relatively conserved among class A GPCRs, and analogous mutations within other GPCRs similarly impaired β-arrestin recruitment while maintaining G protein signaling. To investigate the mechanism of this signaling bias, we used an active-state structure of the β2-adrenergic receptor (β2R) to build β2R-WT and β2R-Y1995.38A models in complex with the full β2R agonist BI-167107 for molecular dynamics simulations. These analyses identified conformational rearrangements in β2R-Y1995.38A that propagated from the extracellular ligand binding site to the intracellular surface, resulting in a modified orientation of the second intracellular loop in β2R-Y1995.38A, which is predicted to affect its interactions with β-arrestin. Our findings provide a structural basis for how ligand binding site alterations can allosterically affect GPCR-transducer interactions and result in biased signaling.

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Sequence, Structure and Ligand Binding Evolution of Rhodopsin-Like G Protein-Coupled Receptors: A Crystal Structure-Based Phylogenetic Analysis

PLoS ONE ◽

10.1371/journal.pone.0123533 ◽

2015 ◽

Vol 10 (4) ◽

pp. e0123533 ◽

Cited By ~ 29

Author(s):

Steffen Wolf ◽

Stefan Grünewald

Keyword(s):

Crystal Structure ◽

Phylogenetic Analysis ◽

Ligand Binding ◽

G Protein ◽

G Protein Coupled Receptors ◽

Sequence Structure ◽

G Protein Coupled

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Multiparametric Homogeneous Method for Identification of Ligand Binding to G Protein-Coupled Receptors: Receptor–Ligand Binding and β-Arrestin Assay

Analytical Chemistry ◽

10.1021/ac303215r ◽

2013 ◽

Vol 85 (4) ◽

pp. 2276-2281 ◽

Cited By ~ 7

Author(s):

Kari Kopra ◽

Markus Kainulainen ◽

Piia Mikkonen ◽

Anita Rozwandowicz-Jansen ◽

Pekka Hänninen ◽

...

Keyword(s):

Ligand Binding ◽

G Protein ◽

G Protein Coupled Receptors ◽

Receptor Ligand ◽

Homogeneous Method ◽

G Protein Coupled

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Family Resemblances? Ligand Binding and Activation of Family A and B G-Protein-Coupled Receptors

Biochemical Society Transactions ◽

10.1042/bst0350707 ◽

2007 ◽

Vol 35 (4) ◽

pp. 707-708 ◽

Cited By ~ 3

Author(s):

D.R. Poyner ◽

M. Wheatley

Keyword(s):

Ligand Binding ◽

G Protein ◽

Present Article ◽

G Protein Coupled Receptors ◽

Society Meeting ◽

Family Resemblances ◽

G Protein Coupled ◽

Compare And Contrast ◽

Present Meeting

In April 2007, the Biochemical Society held a meeting to compare and contrast ligand binding and activation of Family A and B GPCRs (G-protein-coupled receptors). Being the largest class, Family A GPCRs usually receive the most attention, although a previous Biochemical Society meeting has focused on Family B GPCRs. The aim of the present meeting was to bring researchers of both families together in order to identify commonalities between the two. The present article introduces the proceedings of the meeting, briefly commenting on the focus of each of the following articles.

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Locating ligand binding sites in G-protein coupled receptors using combined information from docking and sequence conservation

10.1101/461681 ◽

2018 ◽

Cited By ~ 1

Author(s):

Ashley R. Vidad ◽

Stephen Macaspac ◽

Ho-Leung Ng

Keyword(s):

Ligand Binding ◽

G Protein ◽

Binding Sites ◽

G Protein Coupled Receptors ◽

Sequence Conservation ◽

Surface Geometry ◽

A2a Adenosine Receptors ◽

Ligand Binding Sites ◽

G Protein Coupled ◽

Homology Models

AbstractG-protein coupled receptors (GPCRs) are the largest protein family of drug targets. Detailed mechanisms of binding are unknown for many important GPCR-ligand pairs due to the difficulties of GPCR recombinant expression, biochemistry, and crystallography. We describe our new method, ConDock, for predicting ligand binding sites in GPCRs using combined information from surface conservation and docking starting from crystal structures or homology models. We demonstrate the effectiveness of ConDock on well-characterized GPCRs such as the β2 adrenergic and A2A adenosine receptors. We also demonstrate that ConDock successfully predicts ligand binding sites from high-quality homology models. Finally, we apply ConDock to predict ligand binding sites on a structurally uncharacterized GPCR, GPER. GPER is the G-protein coupled estrogen receptor, with four known ligands: estradiol, G1, G15, and tamoxifen. ConDock predicts that all four ligands bind to the same location on GPER, centered on L119, H307, and N310; this site is deeper in the receptor cleft than predicted by previous studies. We compare the sites predicted by ConDock and traditional methods that utilize information from surface geometry, surface conservation, and ligand chemical interactions. Incorporating sequence conservation information in ConDock overcomes errors introduced from physics-based scoring functions and homology modeling.

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