title: Regulation of adenylyl cyclases by a region outside the minimally functional cytoplasmic domains.
abstract: The highly conserved topological structure of G protein-activated adenylyl cyclases seems unnecessary because the soluble cytoplasmic domains retain regulatory and catalytic properties. Yet, we previously isolated a constitutively active mutant of the Dictyostelium discoideum adenylyl cyclase harboring a single point mutation in the region linking the cytoplasmic and membrane domains (Leu-394). We show here that multiple amino acid substitutions at Leu-394 also display constitutive activity. The constitutive activity of these mutants is not dependent on G proteins or cytosolic regulators, although some of the mutants can be activated to higher levels than wild type. Combining a constitutive mutation such as L394T with K482N, a point mutation that renders the enzyme insensitive to regulators, restores an enzyme with wild type properties of low basal activity and the capacity to be activated by G proteins. Thus regions located outside the cytoplasmic loops of adenylyl cyclases are not only important in the acquisition of an activated conformation, they also have impact on other regions within the catalytic core of the enzyme.
authors.affiliation: Department of Biological Chemistry, The Johns Hopkins School of Medicine, Baltimore, Maryland 21205, USA. parentc@helix.nih.gov
authors.forename: Carole A
authors.initials: CA
authors.lastname: Parent
authors.forename: Jane
authors.initials: J
authors.lastname: Borleis
authors.forename: Peter N
authors.initials: PN
authors.lastname: Devreotes
mesh_terms: D000262:Adenylyl Cyclases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D004023:Dictyostelium / Q000201:enzymology* / Q000235:genetics; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010641:Phenotype; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment
chemical_list: D016244:Guanosine 5'-O-(3-Thiotriphosphate); D000262:Adenylyl Cyclases