title: The unique nature of the serine interactions for alpha 1-adrenergic receptor agonist binding and activation.
abstract: Activation of the beta2- and alpha2-adrenergic receptors (AR) involves hydrogen bonding of serine residues in the fifth transmembrane segment (TMV) to the catechol hydroxyls of the endogenous agonists, epinephrine and norepinephrine. With the beta2-AR both Ser204 and Ser207 but not a third TMV serine (Ser203) are required for binding and full agonist activity. However, with the alpha2a-AR only one of two TMV serines (Ser204, equivalent to Ser207 in the beta-AR) appears to contribute partially to agonist-binding and activation. Because the alpha1a-AR uniquely contains only two TMV serines, this subtype was used to systematically evaluate the role of hydrogen bonding in alpha1-AR activation. Binding of epinephrine or its monohydroxyl congeners, phenylephrine and synephrine, was not decreased when tested with alanine- substitution mutants that lacked either Ser188 (Ser188--> Ala) or Ser192 (Ser192-->Ala). With the substitution of both serines in the double mutant, Ser188/192-->Ala, binding of all three ligands was significantly reduced (10- 100-fold) consistent with a single hydrogen bond interaction. However, receptor-mediated inositol phosphate production was markedly attenuated only with the Ser188-->Ala mutation and not with Ser192-->Ala. In support of the importance of Ser188, binding of phenylephrine (meta-hydroxyl only) by Ser192-->Ala increased 7-fold over that observed with either the wild type receptor or the Ser188-->Ala mutation. Binding of synephrine (para-hydroxyl only) was unchanged with the Ser192-->Ala mutation. In addition, when combined with a recently described constitutively active alpha1a-AR mutation (Met292-->Leu), only the Ser188-->Ala mutation and not Ser192-->Ala relieved the high affinity binding and increased agonist potency observed with the Met292-->Leu mutation. A simple interpretation of these findings is that the meta-hydroxyl of the endogenous agonists preferentially binds to Ser188, and it is this hydrogen bond interaction, and not that between the para-hydroxyl and Ser192, that allows receptor activation. Furthermore, since Ser188 and Ser192 are separated by three residues on the TMV alpha-helix, whereas Ser204 and Ser207 of the beta2-AR are separated by only two residues, the orientation of the catechol ring in the alpha1-AR binding pocket appears to be unique and rotated approximately 120 degrees to that in the beta2-AR.
authors.affiliation: Department of Molecular Cardiology, Cleveland Clinic Research Institute, Ohio 44195, USA.
authors.forename: J
authors.initials: J
authors.lastname: Hwa
authors.forename: D M
authors.initials: DM
authors.lastname: Perez
mesh_terms: D000316:Adrenergic alpha-Agonists / Q000737:chemistry / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D001667:Binding, Competitive; D006224:Cricetinae; D004837:Epinephrine / Q000378:metabolism; D006801:Humans; D006860:Hydrogen Bonding; D008024:Ligands; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010656:Phenylephrine / Q000378:metabolism; D051381:Rats; D018340:Receptors, Adrenergic, alpha-1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018341:Receptors, Adrenergic, alpha-2 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D018343:Receptors, Adrenergic, beta-2 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012694:Serine / Q000737:chemistry; D013578:Synephrine / Q000378:metabolism
chemical_list: D000316:Adrenergic alpha-Agonists; D008024:Ligands; D018340:Receptors, Adrenergic, alpha-1; D018341:Receptors, Adrenergic, alpha-2; D018343:Receptors, Adrenergic, beta-2; D010656:Phenylephrine; D012694:Serine; D013578:Synephrine; D004837:Epinephrine