title: Crystal structures of human soluble adenylyl cyclase reveal mechanisms of catalysis and of its activation through bicarbonate.
abstract: cAMP is an evolutionary conserved, prototypic second messenger regulating numerous cellular functions. In mammals, cAMP is synthesized by one of 10 homologous adenylyl cyclases (ACs): nine transmembrane enzymes and one soluble AC (sAC). Among these, only sAC is directly activated by bicarbonate (HCO3(-)); it thereby serves as a cellular sensor for HCO3(-), carbon dioxide (CO2), and pH in physiological functions, such as sperm activation, aqueous humor formation, and metabolic regulation. Here, we describe crystal structures of human sAC catalytic domains in the apo state and in complex with substrate analog, products, and regulators. The activator HCO3(-) binds adjacent to Arg176, which acts as a switch that enables formation of the catalytic cation sites. An anionic inhibitor, 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid, inhibits sAC through binding to the active site entrance, which blocks HCO3(-) activation through steric hindrance and trapping of the Arg176 side chain. Finally, product complexes reveal small, local rearrangements that facilitate catalysis. Our results provide a molecular mechanism for sAC catalysis and cellular HCO3(-) sensing and a basis for targeting this system with drugs.
authors.affiliation: Department of Biochemistry, University of Bayreuth, 95440 Bayreuth, Germany.
authors.forename: Silke
authors.initials: S
authors.lastname: Kleinboelting
authors.forename: Ana
authors.initials: A
authors.lastname: Diaz
authors.forename: Sebastien
authors.initials: S
authors.lastname: Moniot
authors.forename: Joop
authors.initials: J
authors.lastname: van den Heuvel
authors.forename: Michael
authors.initials: M
authors.lastname: Weyand
authors.forename: Lonny R
authors.initials: LR
authors.lastname: Levin
authors.forename: Jochen
authors.initials: J
authors.lastname: Buck
authors.forename: Clemens
authors.initials: C
authors.lastname: Steegborn
mesh_terms: D000262:Adenylyl Cyclases / Q000737:chemistry*; D002384:Catalysis; D003001:Cloning, Molecular; D003460:Crystallization; D004789:Enzyme Activation / Q000235:genetics / Q000502:physiology*; D006801:Humans; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D015398:Signal Transduction / Q000235:genetics*; D017693:Sodium Bicarbonate / Q000378:metabolism*
chemical_list: D017693:Sodium Bicarbonate; C579271:ADCY10 protein, human; D000262:Adenylyl Cyclases
keywords: activation mechanism; bicarbonate signaling; catalytic mechanism; inhibition mechanism