title: Crystal Structure of G Protein-coupled Receptor Kinase 5 in Complex with a Rationally Designed Inhibitor.
abstract: G protein-coupled receptor kinases (GRKs) regulate cell signaling by initiating the desensitization of active G protein-coupled receptors. The two most widely expressed GRKs (GRK2 and GRK5) play a role in cardiovascular disease and thus represent important targets for the development of novel therapeutic drugs. In the course of a GRK2 structure-based drug design campaign, one inhibitor (CCG215022) exhibited nanomolar IC50 values against both GRK2 and GRK5 and good selectivity against other closely related kinases such as GRK1 and PKA. Treatment of murine cardiomyocytes with CCG215022 resulted in significantly increased contractility at 20-fold lower concentrations than paroxetine, an inhibitor with more modest selectivity for GRK2. A 2.4 Å crystal structure of the GRK5·CCG215022 complex was determined and revealed that the inhibitor binds in the active site similarly to its parent compound GSK180736A. As designed, its 2-pyridylmethyl amide side chain occupies the hydrophobic subsite of the active site where it forms three additional hydrogen bonds, including one with the catalytic lysine. The overall conformation of the GRK5 kinase domain is similar to that of a previously determined structure of GRK6 in what is proposed to be its active state, but the C-terminal region of the enzyme adopts a distinct conformation. The kinetic properties of site-directed mutants in this region are consistent with the hypothesis that this novel C-terminal structure is representative of the membrane-bound conformation of the enzyme.
authors.affiliation: Life Sciences Institute and the Departments of Pharmacology and Biological Sciences, University of Michigan, Ann Arbor, Michigan 48109.
authors.forename: Kristoff T
authors.initials: KT
authors.lastname: Homan
authors.affiliation: Vahlteich Medicinal Chemistry Core and the Department of Medicinal Chemistry, University of Michigan, Ann Arbor, Michigan 48109.
authors.forename: Helen V
authors.initials: HV
authors.lastname: Waldschmidt
authors.affiliation: Life Sciences Institute and the Departments of Pharmacology and Biological Sciences, University of Michigan, Ann Arbor, Michigan 48109.
authors.forename: Alisa
authors.initials: A
authors.lastname: Glukhova
authors.affiliation: Center for Translational Medicine, Temple University School of Medicine, Philadelphia, Pennsylvania 19140.
authors.forename: Alessandro
authors.initials: A
authors.lastname: Cannavo
authors.affiliation: Center for Translational Medicine, Temple University School of Medicine, Philadelphia, Pennsylvania 19140.
authors.forename: Jianliang
authors.initials: J
authors.lastname: Song
authors.affiliation: Center for Translational Medicine, Temple University School of Medicine, Philadelphia, Pennsylvania 19140.
authors.forename: Joseph Y
authors.initials: JY
authors.lastname: Cheung
authors.affiliation: Center for Translational Medicine, Temple University School of Medicine, Philadelphia, Pennsylvania 19140.
authors.forename: Walter J
authors.initials: WJ
authors.lastname: Koch
authors.affiliation: Vahlteich Medicinal Chemistry Core and the Department of Medicinal Chemistry, University of Michigan, Ann Arbor, Michigan 48109.
authors.forename: Scott D
authors.initials: SD
authors.lastname: Larsen
authors.affiliation: Life Sciences Institute and the Departments of Pharmacology and Biological Sciences, University of Michigan, Ann Arbor, Michigan 48109. Electronic address: tesmerjj@umich.edu.
authors.forename: John J G
authors.initials: JJG
authors.lastname: Tesmer
mesh_terms: D000818:Animals; D002317:Cardiovascular Agents / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology; D020134:Catalytic Domain; D002417:Cattle; D018360:Crystallography, X-Ray; D015195:Drug Design; D004791:Enzyme Inhibitors / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology; D054774:G-Protein-Coupled Receptor Kinase 5 / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D015870:Gene Expression; D006346:Heart Septum / Q000737:chemistry / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D006352:Heart Ventricles / Q000737:chemistry / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D006860:Hydrogen Bonding; D057927:Hydrophobic and Hydrophilic Interactions; D007700:Kinetics; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D009200:Myocardial Contraction / Q000187:drug effects; D032383:Myocytes, Cardiac / Q000737:chemistry / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology; D017374:Paroxetine / Q000737:chemistry / Q000494:pharmacology; D054730:Protein Interaction Domains and Motifs; D017433:Protein Structure, Secondary; D011725:Pyridines / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology; D016415:Sequence Alignment
chemical_list: C000602816:CCG215022; D002317:Cardiovascular Agents; D004791:Enzyme Inhibitors; D011725:Pyridines; D017374:Paroxetine; D054774:G-Protein-Coupled Receptor Kinase 5
keywords: G protein-coupled receptor kinase; crystallography; enzyme inhibitor; membrane targeting; plasma membrane; protein kinase; rational drug design; site-directed mutagenesis; x-ray crystallography