title: Deletion within the amino-terminal region of Gs alpha impairs its ability to interact with beta gamma subunits and to activate adenylate cyclase.
abstract: Proteolytic experiments performed on transducin and Go alpha subunit strongly suggest that the amino-terminal residues of the alpha chain are involved in the interaction with beta gamma subunits. To test the possibility that the same region in Gs may fulfill a similar function, we introduced a deletion in the amino-terminal domain of Gs alpha. The properties of the wild type and the deleted alpha chains were characterized on in vitro translated proteins or after reconstitution of cyc- membranes by in vitro-translated alpha subunits. The mutant (delta 2-29) Gs alpha could still bind guanosine 5'-3-O-(thio)triphosphate, as revealed by its resistance to trypsin proteolysis and was still able to interact with the membrane. However, (delta 2-29) Gs alpha was not ADP-ribosylated by cholera toxin. In contrast to Gs alpha, addition of beta gamma subunits did not increase the rate of sedimentation of (delta 2-29) Gs alpha in sucrose gradients. Binding experiments on reconstituted membranes showed that the coupling to beta-adrenergic receptors was very low with (delta 2-29) Gs alpha. Finally, the mutant did not restore activation of adenylate cyclase of cyc- membranes. We propose that the primary functional defect is the loss of interaction with beta gamma subunits, which secondarily impairs beta gamma-dependent properties such as receptor coupling and cholera toxin-catalyzed ADP-ribosylation. However, it remains to be established that the lack of adenylate cyclase activation also results from this impaired interaction with beta gamma subunits.
authors.affiliation: Centre National de la Recherche Scientifique-Institut National de la Santé et de la Recherche Médicale de Pharmacologie et Endocrinologie, Montpellier, France.
authors.forename: L
authors.initials: L
authors.lastname: Journot
authors.forename: C
authors.initials: C
authors.lastname: Pantaloni
authors.forename: J
authors.initials: J
authors.lastname: Bockaert
authors.forename: Y
authors.initials: Y
authors.lastname: Audigier
mesh_terms: D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D002772:Cholera Toxin / Q000494:pharmacology; D004252:DNA Mutational Analysis; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000235:genetics / Q000502:physiology*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D066298:In Vitro Techniques; D007545:Isoproterenol / Q000378:metabolism; D046911:Macromolecular Substances; D051379:Mice; D010446:Peptide Fragments / Q000737:chemistry; D011943:Receptors, Adrenergic, beta / Q000378:metabolism; D011994:Recombinant Proteins; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D014357:Trypsin / Q000494:pharmacology
chemical_list: D046911:Macromolecular Substances; D010446:Peptide Fragments; D011943:Receptors, Adrenergic, beta; D011994:Recombinant Proteins; D000246:Adenosine Diphosphate Ribose; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D002772:Cholera Toxin; D014357:Trypsin; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases; D007545:Isoproterenol