title: Stereochemistry of glutamate receptor agonist efficacy: engineering a dual-specificity AMPA/kainate receptor.
abstract: Upon agonist binding, the bilobate ligand-binding domains of the ionotropic glutamate receptors (iGluR) undergo a cleft closure whose magnitude correlates broadly with the efficacy of the agonist. AMPA (alpha-amino-5-methyl-3-hydroxy-4-isoxazolepropionic acid) and kainate are nonphysiological agonists that distinguish between subsets of iGluR. Kainate acts with low efficacy at AMPA receptors. Here we report that the structure-based mutation L651V converts the GluR4 AMPA receptor into a dual-specificity AMPA/kainate receptor fully activated by both agonists. To probe the stereochemical basis of partial agonism, we have also investigated the correlation between agonist efficacy and a series of vibrational and fluorescence spectroscopic signals of agonist binding to the corresponding wild-type and mutant GluR4 ligand-binding domains. Two signals track the extent of channel activation: the maximal change in intrinsic tryptophan fluorescence and the environment of the single non-disulfide bonded C426, which appears to probe the strength of interactions with the ligand alpha-amino group. Both of these signals arise from functional groups that are poised to detect changes in the extent of channel cleft closure and thus provide additional information about the coupling between conformational changes in the ligand-binding domain and activation of the intact receptor.
authors.affiliation: Department of Biochemistry, Dartmouth Medical School, Hanover, New Hampshire 03755, USA.
authors.forename: Dean R
authors.initials: DR
authors.lastname: Madden
authors.forename: Qing
authors.initials: Q
authors.lastname: Cheng
authors.forename: Shalita
authors.initials: S
authors.lastname: Thiran
authors.forename: Shanti
authors.initials: S
authors.lastname: Rajan
authors.forename: Frank
authors.initials: F
authors.lastname: Rigo
authors.forename: Kari
authors.initials: K
authors.lastname: Keinänen
authors.forename: Stefan
authors.initials: S
authors.lastname: Reinelt
authors.forename: Herbert
authors.initials: H
authors.lastname: Zimmermann
authors.forename: Vasanthi
authors.initials: V
authors.lastname: Jayaraman
mesh_terms: D002460:Cell Line; D004553:Electric Conductivity; D018690:Excitatory Amino Acid Agonists / Q000737:chemistry / Q000494:pharmacology; D006801:Humans; D007608:Kainic Acid / Q000737:chemistry / Q000494:pharmacology; D009154:Mutation; D015202:Protein Engineering; D018091:Receptors, AMPA / Q000819:agonists* / Q000235:genetics* / Q000502:physiology; D018092:Kainic Acid Receptors / Q000819:agonists*; D018350:alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid / Q000737:chemistry / Q000494:pharmacology
chemical_list: D018690:Excitatory Amino Acid Agonists; D007608:Kainic Acid; D018091:Receptors, AMPA; D018092:Kainic Acid Receptors; D018350:alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; C104724:glutamate receptor ionotropic, AMPA 4