title: GPCRs through the keyhole: the role of protein flexibility in ligand binding to β-adrenoceptors.
abstract: G protein-coupled receptors (GPCRs) are proteins of pharmaceutical importance, with over 30% of all drugs in clinical use targeting them. Increasing numbers of X-ray crystal (XRC) structures of GPCRs offer a wealth of data relating to ligand binding. For the β-adrenoceptors (β-ARs), XRC structures are available for human β2- and turkey β1-subtypes, in complexes with a range of ligands. While these structures provide insight into the origins of ligand structure-activity relationships (SARs), questions remain. The ligands in all published complexed XRC structures lack extensive substitution, with no obvious way the ligand-binding site can accommodate β1-AR-selective antagonists with extended side-chains para- to the common aryloxypropanolamine pharmacophore. Using standard computational docking tools with such ligands generally returns poses that fail to explain known SARs. Application of our Active Site Pressurisation modelling method to β-AR XRC structures and homology models, however, reveals a dynamic area in the ligand-binding pocket that, through minor changes in amino acid side chain orientations, opens a fissure between transmembrane helices H4 and H5, exposing intra-membrane space. This fissure, which we term the "keyhole", is ideally located to accommodate extended moieties present in many high-affinity β1-AR-selective ligands, allowing the rest of the ligand structure to adopt a canonical pose in the orthosteric binding site. We propose the keyhole may be a feature of both β1- and β2-ARs, but that subtle structural differences exist between the two, contributing to subtype-selectivity. This has consequences for the rational design of future generations of subtype-selective ligands for these therapeutically important targets.
authors.affiliation: a School of Pharmacy and Centre for Biomolecular Sciences , The University of Nottingham , University Park, Nottingham NG7 2RD , UK.
authors.forename: Abigail L
authors.identifier: http://orcid.org/0000-0001-7763-0076
authors.initials: AL
authors.lastname: Emtage
authors.affiliation: a School of Pharmacy and Centre for Biomolecular Sciences , The University of Nottingham , University Park, Nottingham NG7 2RD , UK.
authors.forename: Shailesh N
authors.identifier: http://orcid.org/0000-0002-2252-1689
authors.initials: SN
authors.lastname: Mistry
authors.affiliation: a School of Pharmacy and Centre for Biomolecular Sciences , The University of Nottingham , University Park, Nottingham NG7 2RD , UK.
authors.forename: Peter M
authors.identifier: http://orcid.org/0000-0002-5866-9271
authors.initials: PM
authors.lastname: Fischer
authors.affiliation: a School of Pharmacy and Centre for Biomolecular Sciences , The University of Nottingham , University Park, Nottingham NG7 2RD , UK.
authors.forename: Barrie
authors.identifier: http://orcid.org/0000-0003-0030-9908
authors.initials: B
authors.lastname: Kellam
authors.affiliation: a School of Pharmacy and Centre for Biomolecular Sciences , The University of Nottingham , University Park, Nottingham NG7 2RD , UK.
authors.forename: Charles A
authors.identifier: http://orcid.org/0000-0003-4090-3960
authors.initials: CA
authors.lastname: Laughton
mesh_terms: D001665:Binding Sites; D018360:Crystallography, X-Ray; D006801:Humans; D008024:Ligands; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D011485:Protein Binding; D011487:Protein Conformation; D018342:Receptors, Adrenergic, beta-1 / Q000737:chemistry / Q000378:metabolism*; D018343:Receptors, Adrenergic, beta-2 / Q000737:chemistry / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled / Q000737:chemistry / Q000378:metabolism*; D013329:Structure-Activity Relationship
chemical_list: D008024:Ligands; D018342:Receptors, Adrenergic, beta-1; D018343:Receptors, Adrenergic, beta-2; D043562:Receptors, G-Protein-Coupled
keywords: GPCRs; active site pressurisation; beta adrenergic receptor; docking; modelling; molecular dynamics; protein flexibility