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

This is the authors' abstract. We don't add key points for this paper.

A conserved amino acid residue in the ligand binding site of class A GPCRs regulates receptor interactions with β-arrestin. A biasing position for GPCRs GPCRs are the largest class of druggable receptors in the human proteome. Drugs that preferentially activate G protein– or β-arrestin–dependent signaling downstream of GPCRs are less likely to come with unwanted side effects. Using biochemical analyses, Sanchez-Soto et al. identified a specific conserved residue in the ligand binding site for multiple GPCRs that modulate β-arrestin–dependent signaling while minimally affecting that mediated by G proteins. Molecular dynamics simulations showed that mutations in this residue resulted in conformational changes that were expected to allosterically affect the interaction of the receptor with β-arrestin. These findings describe a mechanism by which changes in the ligand binding site of GPCRs can result in biased downstream signaling. 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.