Biomedical subjects

Matthew T Geballe

Publications and source records attributed to Matthew T Geballe.

4 recordsLinked to original sources

PubMed2006

Glutamate receptors: variation in structure-function coupling.

Fast excitatory synaptic transmission in the CNS relies almost entirely on the neurotransmitter glutamate and its family of ion channel receptors. An appreciation of the coupling between agonist binding and channel opening has advanced rapidly during the past five years, largely as a result of new structural information about the agonist-binding site. Recent studies suggest that despite many structural similarities different family members use different mechanisms to translate agonist binding into channel opening.

Animals↗

PubMed2005

Mechanism of partial agonism at NMDA receptors for a conformationally restricted glutamate analog.

The NMDA ionotropic glutamate receptor is ubiquitous in mammalian central neurons. Because partial agonists bind to the same site as glutamate but induce less channel activation, these compounds provide an opportunity to probe the mechanism of activation of NMDA-type glutamate receptors. Molecular dynamics simulations and site-directed mutagenesis demonstrate that the partial agonist homoquinolinate interacts differently with binding pocket residues than glutamate. Homoquinolinate and glutamate induce distinct changes in the binding pocket, and the binding pocket exhibits significantly more motion with homoquinolinate bound than with glutamate. Patch-clamp recording demonstrates that single-channel activity induced by glutamate or by homoquinolinate has identical single-channel current amplitude and mean open-channel duration but that homoquinolinate slows activation of channel opening relative to glutamate. We hypothesize that agonist-induced conformational changes in the binding pocket control the efficacy of a subunit-specific activation step that precedes the concerted global change in the receptor-channel complex associated with ion channel opening.

Amino Acid Sequence↗

PubMed2005

A binding site tyrosine shapes desensitization kinetics and agonist potency at GluR2. A mutagenic, kinetic, and crystallographic study.

Binding of an agonist to the 2-amino-3-(3-hydroxy-5-methyl-4-isoxazolyl)-propionic acid (AMPA) receptor family of the glutamate receptors (GluRs) results in rapid activation of an ion channel. Continuous application results in a non-desensitizing response for agonists like kainate, whereas most other agonists, such as the endogenous agonist (S)-glutamate, induce desensitization. We demonstrate that a highly conserved tyrosine, forming a wedge between the agonist and the N-terminal part of the bi-lobed ligand-binding site, plays a key role in the receptor kinetics as well as agonist potency and selectivity. The AMPA receptor GluR2, with mutations in Tyr-450, were expressed in Xenopus laevis oocytes and characterized in a two-electrode voltage clamp setup. The mutation GluR2(Y450A) renders the receptor highly kainate selective, and rapid application of kainate to outside-out patches induced strongly desensitizing currents. When Tyr-450 was substituted with the larger tryptophan, the (S)-glutamate desensitization is attenuated with a 10-fold increase in steady-state/peak currents (19% compared with 1.9% at the wild type). Furthermore, the tryptophan mutant was introduced into the GluR2-S1S2J ligand binding core construct and co-crystallized with kainate, and the 2.1-A x-ray structure revealed a slightly more closed ligand binding core as compared with the wild-type complex. Through genetic manipulations combined with structural and electrophysiological analysis, we report that mutations in position 450 invert the potency of two central agonists while concurrently strongly shaping the agonist efficacy and the desensitization kinetics of the AMPA receptor GluR2.

Alanine↗

PubMed2005

Structural features of the glutamate binding site in recombinant NR1/NR2A N-methyl-D-aspartate receptors determined by site-directed mutagenesis and molecular modeling.

We have used site-directed mutagenesis of amino acids located within the S1 and S2 ligand binding domains of the NR2A N-methyl-D-aspartate (NMDA) receptor subunit to explore the nature of ligand binding. Wild-type or mutated NR1/NR2A NMDA receptors were expressed in Xenopus laevis oocytes and studied using two electrode voltage clamp. We investigated the effects of mutations in the S1 and S2 regions on the potencies of the agonists L-glutamate, L-aspartate, (R,S)-tetrazol-5yl-glycine, and NMDA. Mutation of each of the corresponding residues found in the NR2A receptor subunit, suggested to be contact residues in the GluR2 alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor subunit, caused a rightward shift in the concentration-response curve for each agonist examined. None of the mutations examined altered the efficacy of glutamate as assessed by methanethiosulfonate ethylammonium potentiation of agonist-evoked currents. In addition, none of the mutations altered the potency of glycine. Homology modeling and molecular dynamics were used to evaluate molecular details of ligand binding of both wild-type and mutant receptors, as well as to explore potential explanations for agonist selectivity between glutamate receptor subtypes. The modeling studies support our interpretation of the mutagenesis data and indicate a similar binding strategy for L-glutamate and NMDA when they occupy the binding site in NMDA receptors, as has been proposed for glutamate binding to the GluR2 AMPA receptor subunit. Furthermore, we offer an explanation as to why "charge conserving" mutations of two residues in the binding pocket result in nonfunctional receptor channels and suggest a contributing molecular determinant for why NMDA is not an agonist at AMPA receptors.

Amino Acid Sequence↗