PT - JOURNAL ARTICLE AU - Victor Jun Yu Lim AU - Richard George William Proudman AU - Stefania Monteleone AU - Peter Kolb AU - Jillian Glenda Baker TI - <strong>The isoleucine at position 118 in transmembrane 2 is responsible for the selectivity of xamoterol, nebivolol and ICI89406 for the human β1-adrenoceptor.</strong> AID - 10.1124/molpharm.122.000583 DP - 2022 Jan 01 TA - Molecular Pharmacology PG - MOLPHARM-AR-2022-000583 4099 - http://molpharm.aspetjournals.org/content/early/2022/11/09/molpharm.122.000583.short 4100 - http://molpharm.aspetjournals.org/content/early/2022/11/09/molpharm.122.000583.full AB - Known off-target interactions frequently cause predictable drug side-effects, e.g. β1-antagonists (used for heart disease) risk β2-mediated bronchospasm. Computer-aided drug design would improve if the structural basis of existing drug selectivity was understood. A mutagenesis approach determined the ligand-amino acid interactions required for β1-selective affinity of xamoterol and nebivolol, followed by computer-based modelling to provide possible structural explanations. 3H-CGP12177 whole cell binding was conducted in CHO cells stably expressing human β1, β2 and chimeric β1/β2-adrenoceptors (ARs). Single point mutations were investigated in transiently transfected cells. Modelling studies involved docking ligands into three-dimensional receptor structures and performing Molecular Dynamics simulations, comparing interaction frequencies between apo and holo structures of β1 and β2-ARs. From these observations, an ICI89406 derivative was investigated that gave further insights into selectivity. Stable cell line studies determined that transmembrane 2 was crucial for the β1-selective affinity of xamoterol and nebivolol. Single point mutations determined that the β1-AR isoleucine (I118) rather than the β2 histidine (H93) explained selectivity. Studies of other β1-ligands found I118 was important for ICI89406 selective affinity but not that for betaxolol, bisoprolol or esmolol. Modelling studies suggested that the interaction energies and solvation of β1-I118 and β2-H93 are factors determining selectivity of xamoterol and ICI89406. ICI89406 without its phenyl group loses its high β1-AR affinity, resulting in the same affinity as for the β2-AR. The human β1-AR residue I118 is crucial for the β1-selective affinity of xamoterol, nebivolol and ICI89406, but not all β1-selective compounds. Significance Statement Some ligands have selective binding affinity for the human β1 versus the β2-adrenoceptor however the molecular / structural reason for this is not known. The transmembrane 2 residue isoleucine I118 is responsible for the selective β1-binding of xamoterol, nebivolol and ICI89406, but does not explain the selective β1-binding of betaxolol, bisoprolol or esmolol. Understanding the structural basis of selectivity is important to improve computer aided ligand design and targeting I118 in β1-adrenoceptors is likely to increase β1-selectivity of drugs.