Abstract
G protein-coupled receptors (GPCRs) represent a large class of transmembrane helical proteins which are involved in numerous physiological signaling pathways and therefore represent crucial pharmacological targets. GPCR function and the action of therapeutic molecules are defined by only a few parameters, including receptor basal activity, ligand affinity, intrinsic efficacy and signal bias. These parameters are encoded in characteristic receptor conformations existing in equilibrium and their populations, which are thus of paramount interest for the understanding of receptor (mal-)functions and rational design of improved therapeutics. To this end, the combination of site-directed spin labeling and EPR spectroscopy, in particular double electron–electron resonance (DEER), is exceedingly valuable as it has access to sub-Angstrom spatial resolution and provides a detailed picture of the number and populations of conformations in equilibrium. This review gives an overview of existing DEER studies on GPCRs with a focus on the delineation of structure/function frameworks, highlighting recent developments in data analysis and visualization. We introduce “conformational efficacy” as a parameter to describe ligand-specific shifts in the conformational equilibrium, taking into account the loose coupling between receptor segments observed for different GPCRs using DEER.
Highlights
G protein-coupled receptors (GPCRs) function as promiscuous and yet highly specific signaling proteins (“allosteric microprocessor” [2]) channeling the binding events of distinct ligands towards different and specific physiological outcomes. This finding has been conceptualized in terms of a conformational selection model: a manifold of distinct receptor conformations coexist in equilibrium, each exhibiting differences in the orientation of structural elements, which range from individual amino acids side chains to secondary and tertiary structures
This concept was initially proposed on the basis of comparative MD simulations on β2AR and rhodopsin in order to explain Gi vs. Gs functional selectivity [51]. This idea has been corroborated by a combination of double electron–electron resonance (DEER) and MD simulations showing that Gi and Gs stabilize distinct receptor conformations [30]. These results suggest that GPCRs and their ligands modulate signaling by a specific equilibrium position between several active conformations, in particular distinct TM6 conformations which are identifiable by DEER
Crystal structures of GPCRs indicate that ICL2 may exist in disordered or helical conformations and may represent an important conformational switch critically involved in G protein binding and/or activation [88,89,90]
Summary
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. GPCRs function as promiscuous and yet highly specific signaling proteins (“allosteric microprocessor” [2]) channeling the binding events of distinct ligands towards different and specific physiological outcomes This finding has been conceptualized in terms of a conformational selection model: a manifold of distinct receptor conformations coexist in equilibrium, each exhibiting differences in the orientation of structural elements, which range from individual amino acids side chains to secondary and tertiary structures. By binding and stabilizing specific receptor conformations, ligands modulate transducer interactions and trigger a characteristic cellular signaling response (Figure 1b). Different ligand classes (such as antagonists, balanced/reference and biased agonists) stabilize distinct GPCR conformations, leading to specific efficacies towards transducers. In order to characterize ligand action, for example during drug design, the structural differences of functionally distinct receptor conformations and their equilibrium populadifferences of functionally distinct receptor conformations and their equilibrium populations need to be determined.
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