title: Pharmacological Analysis and Structure Determination of 7-Methylcyanopindolol-Bound β1-Adrenergic Receptor.
abstract: Comparisons between structures of the β1-adrenergic receptor (AR) bound to either agonists, partial agonists, or weak partial agonists led to the proposal that rotamer changes of Ser(5.46), coupled to a contraction of the binding pocket, are sufficient to increase the probability of receptor activation. (RS)-4-[3-(tert-butylamino)-2-hydroxypropoxy]-1H-indole-2-carbonitrile (cyanopindolol) is a weak partial agonist of β1AR and, based on the hypothesis above, we predicted that the addition of a methyl group to form 4-[(2S)-3-(tert-butylamino)-2-hydroxypropoxy]-7-methyl-1H-indole-2-carbonitrile (7-methylcyanopindolol) would dramatically reduce its efficacy. An eight-step synthesis of 7-methylcyanopindolol was developed and its pharmacology was analyzed. 7-Methylcyanopindolol bound with similar affinity to cyanopindolol to both β1AR and β2AR. As predicted, the efficacy of 7-methylcyanopindolol was reduced significantly compared with cyanopindolol, acting as a very weak partial agonist of turkey β1AR and an inverse agonist of human β2AR. The structure of 7-methylcyanopindolol-bound β1AR was determined to 2.4-Å resolution and found to be virtually identical to the structure of cyanopindolol-bound β1AR. The major differences in the orthosteric binding pocket are that it has expanded by 0.3 Å in 7-methylcyanopindolol-bound β1AR and the hydroxyl group of Ser(5.46) is positioned 0.8 Å further from the ligand, with respect to the position of the Ser(5.46) side chain in cyanopindolol-bound β1AR. Thus, the molecular basis for the reduction in efficacy of 7-methylcyanopindolol compared with cyanopindolol may be regarded as the opposite of the mechanism proposed for the increase in efficacy of agonists compared with antagonists.
authors.affiliation: MRC Laboratory of Molecular Biology, Cambridge Biomedical Campus, Cambridge, United Kingdom (T.S., T.W., A.G.W.L., C.G.T.); Heptares Therapeutics Ltd, Welwyn Garden City, United Kingdom (G.A.B., M.C.); School of Life Sciences, University of Nottingham, Medical School, Queen's Medical Centre, Nottingham, United Kingdom (J.B.); KEK High Energy Accelerator Research Organization, Institute of Materials Structure Science, Structural Biology Research Center, Tsukuba, Japan (T.S.).
authors.forename: Tomomi
authors.initials: T
authors.lastname: Sato
authors.affiliation: MRC Laboratory of Molecular Biology, Cambridge Biomedical Campus, Cambridge, United Kingdom (T.S., T.W., A.G.W.L., C.G.T.); Heptares Therapeutics Ltd, Welwyn Garden City, United Kingdom (G.A.B., M.C.); School of Life Sciences, University of Nottingham, Medical School, Queen's Medical Centre, Nottingham, United Kingdom (J.B.); KEK High Energy Accelerator Research Organization, Institute of Materials Structure Science, Structural Biology Research Center, Tsukuba, Japan (T.S.).
authors.forename: Jillian
authors.initials: J
authors.lastname: Baker
authors.affiliation: MRC Laboratory of Molecular Biology, Cambridge Biomedical Campus, Cambridge, United Kingdom (T.S., T.W., A.G.W.L., C.G.T.); Heptares Therapeutics Ltd, Welwyn Garden City, United Kingdom (G.A.B., M.C.); School of Life Sciences, University of Nottingham, Medical School, Queen's Medical Centre, Nottingham, United Kingdom (J.B.); KEK High Energy Accelerator Research Organization, Institute of Materials Structure Science, Structural Biology Research Center, Tsukuba, Japan (T.S.).
authors.forename: Tony
authors.initials: T
authors.lastname: Warne
authors.affiliation: MRC Laboratory of Molecular Biology, Cambridge Biomedical Campus, Cambridge, United Kingdom (T.S., T.W., A.G.W.L., C.G.T.); Heptares Therapeutics Ltd, Welwyn Garden City, United Kingdom (G.A.B., M.C.); School of Life Sciences, University of Nottingham, Medical School, Queen's Medical Centre, Nottingham, United Kingdom (J.B.); KEK High Energy Accelerator Research Organization, Institute of Materials Structure Science, Structural Biology Research Center, Tsukuba, Japan (T.S.).
authors.forename: Giles A
authors.initials: GA
authors.lastname: Brown
authors.affiliation: MRC Laboratory of Molecular Biology, Cambridge Biomedical Campus, Cambridge, United Kingdom (T.S., T.W., A.G.W.L., C.G.T.); Heptares Therapeutics Ltd, Welwyn Garden City, United Kingdom (G.A.B., M.C.); School of Life Sciences, University of Nottingham, Medical School, Queen's Medical Centre, Nottingham, United Kingdom (J.B.); KEK High Energy Accelerator Research Organization, Institute of Materials Structure Science, Structural Biology Research Center, Tsukuba, Japan (T.S.).
authors.forename: Andrew G W
authors.initials: AG
authors.lastname: Leslie
authors.affiliation: MRC Laboratory of Molecular Biology, Cambridge Biomedical Campus, Cambridge, United Kingdom (T.S., T.W., A.G.W.L., C.G.T.); Heptares Therapeutics Ltd, Welwyn Garden City, United Kingdom (G.A.B., M.C.); School of Life Sciences, University of Nottingham, Medical School, Queen's Medical Centre, Nottingham, United Kingdom (J.B.); KEK High Energy Accelerator Research Organization, Institute of Materials Structure Science, Structural Biology Research Center, Tsukuba, Japan (T.S.).
authors.forename: Miles
authors.initials: M
authors.lastname: Congreve
authors.affiliation: MRC Laboratory of Molecular Biology, Cambridge Biomedical Campus, Cambridge, United Kingdom (T.S., T.W., A.G.W.L., C.G.T.); Heptares Therapeutics Ltd, Welwyn Garden City, United Kingdom (G.A.B., M.C.); School of Life Sciences, University of Nottingham, Medical School, Queen's Medical Centre, Nottingham, United Kingdom (J.B.); KEK High Energy Accelerator Research Organization, Institute of Materials Structure Science, Structural Biology Research Center, Tsukuba, Japan (T.S.) cgt@mrc-lmb.cam.ac.uk.
authors.forename: Christopher G
authors.identifier: http://orcid.org/0000-0002-2008-9183
authors.initials: CG
authors.lastname: Tate
mesh_terms: D000818:Animals; D001665:Binding Sites / Q000502:physiology; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D006801:Humans; D010869:Pindolol / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D011485:Protein Binding / Q000502:physiology; D017433:Protein Structure, Secondary; D018342:Receptors, Adrenergic, beta-1 / Q000737:chemistry* / Q000378:metabolism*; D013329:Structure-Activity Relationship; D014421:Turkey
chemical_list: D018342:Receptors, Adrenergic, beta-1; C047975:cyanopindolol; D010869:Pindolol