Constitutive activation of the alpha 1B-adrenergic receptor by all amino acid substitutions at a single site. Evidence for a region which constrains receptor activation.
Mutations in an intracellular region of the alpha 1B-adrenergic receptor constitutively activate the receptor, resulting in G protein coupling in the absence of agonist, as evidenced by elevated levels of polyphosphoinositide hydrolysis. Remarkably, all 19 possible amino acid substitutions at a single site in this region (alanine 293) confer constitutive activity. This set of mutated receptors exhibits a graded range of elevated biological activities, apparently representing a spectrum of receptor conformations which mimic the "active" state of the wild type receptor. In addition to their constitutive activities, these mutated receptors all demonstrate a higher affinity for agonists, another primary characteristic of the "active" conformation of G protein-coupled receptors. The fact that all possible mutations at this particular site result in increased activity suggests that this region may function to constrain the G protein coupling of the receptor, a constraint which is normally relieved by agonist occupancy.
Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710.
M A
MA
Kjelsberg
S
S
Cotecchia
J
J
Ostrowski
M G
MG
Caron
R J
RJ
Lefkowitz
D000318:Adrenergic beta-Agonists / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004247:DNA / Q000235:genetics; D004837:Epinephrine / Q000378:metabolism / Q000494:pharmacology; D007295:Inositol Phosphates / Q000378:metabolism; D007700:Kinetics; D008961:Models, Structural; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010627:Phenethylamines / Q000378:metabolism; D011487:Protein Conformation; D011943:Receptors, Adrenergic, beta / Q000235:genetics / Q000378:metabolism*; D044024:Tetralones; D014162:Transfection
D000318:Adrenergic beta-Agonists; D007295:Inositol Phosphates; D010627:Phenethylamines; D011943:Receptors, Adrenergic, beta; D044024:Tetralones; D004247:DNA; C048008:2-(beta-(3-iodo-4-hydroxyphenyl)ethylaminomethyl)tetralone; D004837:Epinephrine