Structure of the S1S2 glutamate binding domain of GLuR3

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The ␣-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) subtype of ionotropic glutamate receptors mediates much of the fast excitatory neurotransmission in the central nervous system. The ability of these receptors to shape such responses appears to be due in part to dynamic processes induced by agonists in the ligand-binding domain. Previous studies employing fluorescence spectroscopy and whole cell recording suggest that agonist binding is followed by sequential transitions to one or more distinct conformational states. Here, we used hydrogen-deuterium exchange to determine the mechanisms of binding of glutamate and kainate (full and partial agonists, respectively) to a soluble ligand-binding domain of GluR2. Our results provide a structural basis for sequential state models of agonist binding and the free energy changes of the associated state-to-state transitions. For glutamate, a multi-equilibrium binding reaction was discerned involving distinct ligand docking, domain isomerization, and lobe-locking steps. In contrast, kainate binding involves a simpler dock-isomerization process in which the isomerization equilibrium is shifted dramatically toward open domain conformations. In light of increasing evidence that the stability, in addition to the extent, of domain closure is a critical component of the channel activation mechanism, the differences in domain opening and closing equilibria detected for glutamate and kainate should be useful structural measures for interpreting the markedly different current responses evoked by these agonists.As a consequence of glutamate binding, AMPA 2 receptors produce a rapid flux of cations into post-synaptic neurons that is vital for information transfer in the central nervous system. In addition, these receptors undergo fast deactivation and desensitization along with slower recovery kinetics, which together shape the time course of current decay and restoration (1-6). Structurally, AMPA receptors are tetrameric membranebound proteins composed of modular subunits having two large extracellular domains, a membrane-spanning ion channel, and a C-terminal intracellular region (4, 7-12). One of the extracellular domains, S1S2, is a bilobed structure that binds agonists in a cleft between its lobes (13). Domain closure of S1S2 around the agonist leads to channel opening, and maximum currents have been attributed to complete lobe closure upon complex formation with full agonists (13-17). However, among the full agonists examined, those like glutamate, which are less potent and which bind to S1S2 with lower affinity, produce higher rates of channel deactivation and resensitization (18), despite inducing the same degree of cleft closure in S1S2. This suggests that AMPA receptor function is dependent not only on the extent of lobe closure caused by the agonist but also on the lobe closing and opening dynamics associated with agonist binding and dissociation (19).A prevailing model for the binding reaction is

Section: Discussionmentioning

confidence: 99%

On the Mechanisms of α-Amino-3-hydroxy-5-methylisoxazole-4-propionic Acid (AMPA) Receptor Binding to Glutamate and Kainate

Fenwick

Oswald

2010

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“…In addition, dynamics near the lobe interface suggest that large-scale lobe motions may be present when the LBD is bound to IW (21,23). In contrast, when the wild type LBD is bound to kainate, the lobes are in a relatively open orientation in all crystal structures to date (4,18,24,25), and this is supported by the average structure determined using NMR spectroscopy (15). The idea has been that the lobes remain separated by a steric clash between the isoprenyl group of kainate and the side chain of Leu-650 (3,18).…”

Section: Discussionmentioning

confidence: 99%

“…Although the lobes are slightly open relative to glutamate, two interlobe H-bonds (Gly-451-Gly-653 and Tyr-450-H 2 O-Ser-652) produced by a 180°rota-tion of the peptide bond are present in all three copies. These H-bonds are absent in most crystal structures of partial agonists and have been absent in all previously determined structures of kainate-bound AMPA receptors (3,18,24,25). Previous crystal structures have suggested that kainate prevents further lobe closure due to the steric clash between the side chain of Leu-650 with the isoprenyl group of kainate (foot-in-the-door mechanism) (18).…”

Section: Lobe-locking Mutations-the Ligand-binding Domain Ofmentioning

confidence: 99%

“…Furthermore, little evidence for microseconds-to-milliseconds timescale dynamics in the presence of kainate have been observed in NMR studies (23). However, structures of GluA3 (24) and GluA4 (25) in the presence of kainate are more closed than GluA2 and the D651A mutation of GluA3 results in even further closure of the lobes due to a rotation of the side chain of Leu-650 (GluA2 numbering). 3 Cysteine-trapping studies (i.e.…”

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confidence: 99%

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Mechanism of AMPA Receptor Activation by Partial Agonists

Ahmed

Wang

Chuang

et al. 2011

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The mechanism by which agonist binding to an ionotropic glutamate receptor leads to channel opening is a central issue in molecular neurobiology. Partial agonists are useful tools for studying the activation mechanism because they produce full channel activation with lower probability than full agonists. Structural transitions that determine the efficacy of partial agonists can provide information on the trigger that begins the channel-opening process. The ligand-binding domain of AMPA receptors is a bilobed structure, and the closure of the lobes is associated with channel activation. One possibility is that partial agonists sterically block full lobe closure but that partial degrees of closure trigger the channel with a lower probability. Alternatively, full lobe closure may be required for activation, and the stability of the fully closed state could determine efficacy with the fully closed state having a lower stability when bound to partial relative to full agonists. Disulfide-trapping experiments demonstrated that even extremely low efficacy ligands such as 6-cyano-7-nitroquinoxaline-2,3-dione can produce a full lobe closure, presumably with low probability. The results are consistent the hypothesis that the efficacy is determined at least in part by the stability of the state in which the lobes are fully closed.Ionotropic glutamate receptors are the major mediators of excitatory synaptic transmission in the vertebrate nervous system (1, 2). Glutamate receptor subunits are categorized by pharmacological properties, biological role, and sequence into those that are sensitive to the following: 1) ␣-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA; GluA1-4) 2 ; 2) the neurotoxin kainate (GluK1-5); and 3) N-methyl-D-aspartic acid (NMDA; GluN1, GluN2A-D, GluN3A-B). The important structural details of glutamate receptors were determined first by the solution of the structures of the ligand-binding domain (3-7) and the N-terminal domain (8, 9) followed by the structure of the full-length tetrameric GluA2 subtype of AMPA receptor (10). The structures in the presence of various agonists, partial agonists, and antagonists in combination with spectroscopic measurements, electrophysiological measurements, and site-directed mutagenesis have provided a wealth of information on the link between structure and function (11-13). The binding domain is a bilobed structure to which agonist binds in the cleft between the two lobes. Two linker peptide strands connect the lobes, and the lobes can close to envelope the agonist. One lobe (lobe 1) forms a dimer interface with a second copy of the ligand-binding domain within the tetrameric structure, and the second lobe (lobe 2) is linked to the ion channel domain. When the dimer interface is intact, the force generated by the closing of the lobes can affect the ion channel and presumably open a gate allowing ions to traverse the channel. Complexities arise due to the tetrameric structure and subtle differences between glutamate receptor subtypes, but the general outline of...

“…1). The structure of the nearly fulllength AMPA receptor, GluA2, has been determined (11), and structures of the extracellular domains of all three subtypes have been determined (12)(13)(14)(15)(16)(17). Structures of the LBD of AMPA receptors show that full agonists bind in the cleft between two lobes, and after binding to Lobe 1, the lobes close to allow inter-* This work was supported, in whole or in part, National Institutes of Health Grant R01-GM068935 (to R. E.…”

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confidence: 99%

The Structure of (−)-Kaitocephalin Bound to the Ligand Binding Domain of the (S)-α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid (AMPA)/Glutamate Receptor, GluA2

Ahmed

Hamada

Shinada

et al. 2012

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Background:The natural product (Ϫ)-kaitocephalin is a potential scaffold for development of subtype-specific glutamate receptor antagonists. Results: The crystal structure of (Ϫ)-kaitocephalin bound to an AMPA receptor ligand binding domain was determined. Conclusion: The orientation of (Ϫ)-kaitocephalin in the binding site can explain the subtype selectivity of the parent compound. Significance: Comparisons with the structure of other glutamate receptors provides avenues for development of new subtypespecific antagonists.