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Biased signalling represents the ability of G protein-coupled receptors (GPCRs) to engage distinct pathways with various efficacies depending on the ligand used or on mutations in the receptor. The angiotensin-II type 1 receptor (AT1), a prototypical class A GPCR, can activate various effectors upon stimulation with the endogenous ligand AngII including the Gq/11 protein and β-arrestins. It is believed that the activation of those two pathways can be associated to distinct conformations of the AT1 receptor. To verify this hypothesis, microseconds of molecular dynamics simulations (MD) were computed to explore the conformational landscape sampled by the WT-AT1 receptor, the N111G-AT1 receptor (constitutively active and biased for the Gq/11 pathway) and the D74N-AT1 receptor (biased for the β-arrestin 1 and 2 pathways) in their apo forms and in complex with AngII. The MD simulations of the AngII-WT-AT1, N111G-AT1 and AngII-N111G-AT1 receptors revealed specific structural rearrangements compared to the initial and ground state of the receptor. Simulations of the D74N-AT1 receptor revealed that the mutation stabilizes the receptor in the initial ground state. The presence of AngII further stabilized the ground state of the D74N-AT1 receptor. The biased agonist [Sar¹, Ile8]AngII also showed a preference for the ground state of the WT-AT1 receptor compared to AngII. These results suggest that activation of the Gq/11 pathway is associated with a specific conformational transition stabilized by the agonist whereas the activation of the β-arrestins pathway is linked to the stabilization of the ground state of the receptor. [ABSTRACT FROM AUTHOR]

Breakthroughs in G protein-coupled receptor structure determination based on crystallography have been mainly obtained from receptors occupied in their transmembrane domain core by low molecular weight ligands, and we have only recently begun to elucidate how the extracellular surface of G protein-coupled receptors (GPCRs) allows for the binding of larger peptide molecules. In the present study, we used a unique chemoselective photoaffinity labeling strategy, the methionine proximity assay, to directly identify at physiological conditions a total of 38 discrete ligand/receptor contact residues that form the extracellular peptide-binding site of an activated GPCR, the angiotensin II type 1 receptor. This experimental data set was used in homology modeling to guide the positioning of the angiotensin II (AngII) peptide within several GPCR crystal structure templates. We found that the CXC chemokine receptor type 4 accommodated the results better than the other templates evaluated; ligand/receptor contact residues were spatially grouped into defined interaction clusters with AngII. In the resulting receptor structure, a β-hairpin fold in extracellular loop 2 in conjunction with two extracellular disulfide bridges appeared to open and shape the entrance of the ligand-binding site. The bound AngII adopted a somewhat vertical binding mode, allowing concomitant contacts across the extracellular surface and deep within the transmembrane domain core of the receptor. We propose that such a dualistic nature of GPCR interaction could be well suited for diffusible linear peptide ligands and a common feature of other peptidergic class A GPCRs. [ABSTRACT FROM AUTHOR]

G protein coupled receptors (GPCRs) produce pleiotropic effects by their capacity to engage numerous signaling pathways once activated. Functional selectivity (also called biased signaling), where specific compounds can bring GPCRs to adopt conformations that enable selective receptor coupling to distinct signaling pathways, continues to be significantly investigated. However, an important but often overlooked aspect of functional selectivity is the capability of ligands such as angiotensin II (AngII) to adopt specific conformations that may preferentially bind to selective GPCRs structures. Understanding both receptor and ligand conformation is of the utmost importance for the design of new drugs targeting GPCRs. In this study, we examined the properties of AngII cyclic analogs to impart biased agonism on the angiotensin type 1 receptor (AT 1 R). Positions 3 and 5 of AngII were substituted for cysteine and homocysteine residues ([Sar 1 Hcy 3,5 ]AngII, [Sar 1 Cys 3 Hcy 5 ]AngII and [Sar 1 Cys 3,5 ]AngII) and the resulting analogs were evaluated for their capacity to activate the Gq/11, G12, Gi2, Gi3, Gz, ERK and β-arrestin (βarr) signaling pathways via AT 1 R. Interestingly, [Sar 1 Hcy 3,5 ]AngII exhibited potency and full efficacy on all pathways tested with the exception of the Gq pathway. Molecular dynamic simulations showed that the energy barrier associated with the insertion of residue Phe 8 of AngII within the hydrophobic core of AT 1 R, associated with Gq/11 activation, is increased with [Sar 1 Hcy 3,5 ]AngII. These results suggest that constraining the movements of molecular determinants within a given ligand by introducing cyclic structures may lead to the generation of novel ligands providing more efficient biased agonism., (Copyright © 2018 Elsevier Inc. All rights reserved.)