title: Bithionol Potently Inhibits Human Soluble Adenylyl Cyclase through Binding to the Allosteric Activator Site.
abstract: The signaling molecule cAMP regulates functions ranging from bacterial transcription to mammalian memory. In mammals, cAMP is synthesized by nine transmembrane adenylyl cyclases (ACs) and one soluble AC (sAC). Despite similarities in their catalytic domains, these ACs differ in regulation. Transmembrane ACs respond to G proteins, whereas sAC is uniquely activated by bicarbonate. Via bicarbonate regulation, sAC acts as a physiological sensor for pH/bicarbonate/CO2, and it has been implicated as a therapeutic target, e.g. for diabetes, glaucoma, and a male contraceptive. Here we identify the bisphenols bithionol and hexachlorophene as potent, sAC-specific inhibitors. Inhibition appears mostly non-competitive with the substrate ATP, indicating that they act via an allosteric site. To analyze the interaction details, we solved a crystal structure of an sAC·bithionol complex. The structure reveals that the compounds are selective for sAC because they bind to the sAC-specific, allosteric binding site for the physiological activator bicarbonate. Structural comparison of the bithionol complex with apo-sAC and other sAC·ligand complexes along with mutagenesis experiments reveals an allosteric mechanism of inhibition; the compound induces rearrangements of substrate binding residues and of Arg(176), a trigger between the active site and allosteric site. Our results thus provide 1) novel insights into the communication between allosteric regulatory and active sites, 2) a novel mechanism for sAC inhibition, and 3) pharmacological compounds targeting this allosteric site and utilizing this mode of inhibition. These studies provide support for the future development of sAC-modulating drugs.
authors.affiliation: From the Department of Biochemistry, University of Bayreuth, 95440 Bayreuth, Germany.
authors.forename: Silke
authors.initials: S
authors.lastname: Kleinboelting
authors.affiliation: Department of Pharmacology, Weill Cornell Medical College, New York, New York 10065, High Throughput Screening and Spectroscopy Resource Center, The Rockefeller University, New York, New York 10065, and.
authors.forename: Lavoisier
authors.initials: L
authors.lastname: Ramos-Espiritu
authors.affiliation: Department of Pharmacology, Weill Cornell Medical College, New York, New York 10065.
authors.forename: Hannes
authors.initials: H
authors.lastname: Buck
authors.affiliation: Department of Pharmacology, Weill Cornell Medical College, New York, New York 10065.
authors.forename: Laureen
authors.initials: L
authors.lastname: Colis
authors.affiliation: Helmholtz-Zentrum für Infektionsforschung, 38124 Braunschweig, Germany.
authors.forename: Joop
authors.initials: J
authors.lastname: van den Heuvel
authors.affiliation: High Throughput Screening and Spectroscopy Resource Center, The Rockefeller University, New York, New York 10065, and.
authors.forename: J Fraser
authors.initials: JF
authors.lastname: Glickman
authors.affiliation: Department of Pharmacology, Weill Cornell Medical College, New York, New York 10065.
authors.forename: Lonny R
authors.initials: LR
authors.lastname: Levin
authors.affiliation: Department of Pharmacology, Weill Cornell Medical College, New York, New York 10065.
authors.forename: Jochen
authors.initials: J
authors.lastname: Buck
authors.affiliation: From the Department of Biochemistry, University of Bayreuth, 95440 Bayreuth, Germany, clemens.steegborn@uni-bayreuth.de.
authors.forename: Clemens
authors.initials: C
authors.lastname: Steegborn
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry*; D000262:Adenylyl Cyclases / Q000737:chemistry*; D000494:Allosteric Regulation; D001639:Bicarbonates / Q000737:chemistry*; D001735:Bithionol / Q000737:chemistry*; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D006801:Humans
chemical_list: D001639:Bicarbonates; D000255:Adenosine Triphosphate; D001735:Bithionol; C579271:ADCY10 protein, human; D000262:Adenylyl Cyclases
keywords: adenylate cyclase (adenylyl cyclase); allosteric regulation; crystal structure; cyclic AMP (cAMP); inhibitor