Ligand Binding Ensembles Determine Graded Agonist Efficacies at a G Protein-coupled Receptor.
G protein-coupled receptors constitute the largest family of membrane receptors and modulate almost every physiological process in humans. Binding of agonists to G protein-coupled receptors induces a shift from inactive to active receptor conformations. Biophysical studies of the dynamic equilibrium of receptors suggest that a portion of receptors can remain in inactive states even in the presence of saturating concentrations of agonist and G protein mimetic. However, the molecular details of agonist-bound inactive receptors are poorly understood. Here we use the model of bitopic orthosteric/allosteric (i.e. dualsteric) agonists for muscarinic M2 receptors to demonstrate the existence and function of such inactive agonist·receptor complexes on a molecular level. Using all-atom molecular dynamics simulations, dynophores (i.e. a combination of static three-dimensional pharmacophores and molecular dynamics-based conformational sampling), ligand design, and receptor mutagenesis, we show that inactive agonist·receptor complexes can result from agonist binding to the allosteric vestibule alone, whereas the dualsteric binding mode produces active receptors. Each agonist forms a distinct ligand binding ensemble, and different agonist efficacies depend on the fraction of purely allosteric (i.e. inactive) versus dualsteric (i.e. active) binding modes. We propose that this concept may explain why agonist·receptor complexes can be inactive and that adopting multiple binding modes may be generalized also to small agonists where binding modes will be only subtly different and confined to only one binding site.
From the Institute of Pharmacology and Toxicology, University of Würzburg, Versbacher Strasse 9, 97078 Würzburg, Germany, andreas.bock@uni-wuerzburg.de.
Andreas
A
Bock
Institute of Pharmacy, Freie Universität Berlin, Königin-Luise-Strasse 2 und 4, 14195 Berlin, Germany, m.bermudez@fu-berlin.de.
Marcel
M
Bermudez
Pharmacology and Toxicology Section, Institute of Pharmacy, University of Bonn, Gerhard-Domagk-Strasse 3, 53121 Bonn, Germany.
Fabian
F
Krebs
Dipartimento di Scienze Farmaceutiche, Sezione di Chimica Farmaceutica "Pietro Pratesi," Università degli Studi di Milano, Via Mangiagalli 25, 20133 Milan, Italy, and.
Carlo
C
Matera
Pharmacology and Toxicology Section, Institute of Pharmacy, University of Bonn, Gerhard-Domagk-Strasse 3, 53121 Bonn, Germany.
Brian
B
Chirinda
Institute of Pharmacy, Freie Universität Berlin, Königin-Luise-Strasse 2 und 4, 14195 Berlin, Germany.
Dominique
D
Sydow
Dipartimento di Scienze Farmaceutiche, Sezione di Chimica Farmaceutica "Pietro Pratesi," Università degli Studi di Milano, Via Mangiagalli 25, 20133 Milan, Italy, and.
Clelia
C
Dallanoce
Institute of Pharmacy, University of Würzburg, Am Hubland, 97074 Würzburg, Germany.
Ulrike
U
Holzgrabe
Dipartimento di Scienze Farmaceutiche, Sezione di Chimica Farmaceutica "Pietro Pratesi," Università degli Studi di Milano, Via Mangiagalli 25, 20133 Milan, Italy, and.
Marco
M
De Amici
From the Institute of Pharmacology and Toxicology, University of Würzburg, Versbacher Strasse 9, 97078 Würzburg, Germany.
Martin J
MJ
Lohse
Institute of Pharmacy, Freie Universität Berlin, Königin-Luise-Strasse 2 und 4, 14195 Berlin, Germany.
Gerhard
G
Wolber
Pharmacology and Toxicology Section, Institute of Pharmacy, University of Bonn, Gerhard-Domagk-Strasse 3, 53121 Bonn, Germany, k.mohr@uni-bonn.de.
Klaus
K
Mohr
D000494:Allosteric Regulation; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D006801:Humans; D008024:Ligands; D056004:Molecular Dynamics Simulation; D043585:Receptor, Muscarinic M2 / Q000819:agonists* / Q000737:chemistry* / Q000378:metabolism
D008024:Ligands; D043585:Receptor, Muscarinic M2
G protein-coupled receptor (GPCR); chemical biology; drug design; dualsteric ligands; dynamic ligand binding; dynophores; ligand binding ensembles; molecular dynamics; molecular pharmacology; partial agonist