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Structural mechanism of glutamate receptor activation and desensitization.

Meyerson JR, Kumar J, Chittori S, Rao P, Pierson J, Bartesaghi A, Mayer ML, Subramaniam S - Nature (2014)

Bottom Line: Desensitization is accompanied by disruption of the amino-terminal domain tetramer in AMPA, but not kainate, receptors with a two-fold to four-fold symmetry transition in the ligand-binding domains in both subtypes.The 7.6 Å structure of a desensitized kainate receptor shows how these changes accommodate channel closing.These findings integrate previous physiological, biochemical and structural analyses of glutamate receptors and provide a molecular explanation for key steps in receptor gating.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Cell Biology, Center for Cancer Research, NCI, NIH, Bethesda, Maryland 20892, USA.

ABSTRACT
Ionotropic glutamate receptors are ligand-gated ion channels that mediate excitatory synaptic transmission in the vertebrate brain. To gain a better understanding of how structural changes gate ion flux across the membrane, we trapped rat AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid) and kainate receptor subtypes in their major functional states and analysed the resulting structures using cryo-electron microscopy. We show that transition to the active state involves a 'corkscrew' motion of the receptor assembly, driven by closure of the ligand-binding domain. Desensitization is accompanied by disruption of the amino-terminal domain tetramer in AMPA, but not kainate, receptors with a two-fold to four-fold symmetry transition in the ligand-binding domains in both subtypes. The 7.6 Å structure of a desensitized kainate receptor shows how these changes accommodate channel closing. These findings integrate previous physiological, biochemical and structural analyses of glutamate receptors and provide a molecular explanation for key steps in receptor gating.

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Restored open state density map for the GluA2 Quisqualate complexa, Density map for the GluA2em open state obtained by addition of the allosteric modulator LY451646 to a suspension of quisqualate-bound, desensitized GluA2. The purpose of the experiment was to test whether structural changes resulting from quisqualate binding to generate the desensitized state could be reversed by addition of an excess of the allosteric modulator LY451646, used to stabilize the open state. The map display shown at left is color-coded to highlight variation in resolution across different regions of the map. b, Density map for the glutamate bound open state obtained by addition of LY451646 30 min prior to agonist, as shown in Figure 2. The map display shown at left is color-coded as in (a) to highlight variation in resolution across different regions of the map. Comparison of the two maps and the fits of ATD and LBD dimers shows that they are essentially identical, establishing the conformational changes that occur with desensitization are reversible and can be modulated by allosteric modulators.
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Figure 11: Restored open state density map for the GluA2 Quisqualate complexa, Density map for the GluA2em open state obtained by addition of the allosteric modulator LY451646 to a suspension of quisqualate-bound, desensitized GluA2. The purpose of the experiment was to test whether structural changes resulting from quisqualate binding to generate the desensitized state could be reversed by addition of an excess of the allosteric modulator LY451646, used to stabilize the open state. The map display shown at left is color-coded to highlight variation in resolution across different regions of the map. b, Density map for the glutamate bound open state obtained by addition of LY451646 30 min prior to agonist, as shown in Figure 2. The map display shown at left is color-coded as in (a) to highlight variation in resolution across different regions of the map. Comparison of the two maps and the fits of ATD and LBD dimers shows that they are essentially identical, establishing the conformational changes that occur with desensitization are reversible and can be modulated by allosteric modulators.

Mentions: To determine whether conformational variability observed for quisqualate-GluA2em complexes reflects the properties of functional receptors, we tested whether subsequent addition of 0.5 mM LY451646 to the same preparation used for cryo-EM analysis of the desensitized state would restore a homogeneous active state conformation. Structural analysis at ~ 16 Å resolution of GluA2em receptor suspensions treated this way demonstrates that the active conformation is indeed restored (Fig. 3c and Extended Data Fig. 6). Our experiments thus establish that in the desensitized state, quisqualate-bound GluA2 is fully functional and capable of undergoing conversion to the active state.


Structural mechanism of glutamate receptor activation and desensitization.

Meyerson JR, Kumar J, Chittori S, Rao P, Pierson J, Bartesaghi A, Mayer ML, Subramaniam S - Nature (2014)

Restored open state density map for the GluA2 Quisqualate complexa, Density map for the GluA2em open state obtained by addition of the allosteric modulator LY451646 to a suspension of quisqualate-bound, desensitized GluA2. The purpose of the experiment was to test whether structural changes resulting from quisqualate binding to generate the desensitized state could be reversed by addition of an excess of the allosteric modulator LY451646, used to stabilize the open state. The map display shown at left is color-coded to highlight variation in resolution across different regions of the map. b, Density map for the glutamate bound open state obtained by addition of LY451646 30 min prior to agonist, as shown in Figure 2. The map display shown at left is color-coded as in (a) to highlight variation in resolution across different regions of the map. Comparison of the two maps and the fits of ATD and LBD dimers shows that they are essentially identical, establishing the conformational changes that occur with desensitization are reversible and can be modulated by allosteric modulators.
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Related In: Results  -  Collection

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Figure 11: Restored open state density map for the GluA2 Quisqualate complexa, Density map for the GluA2em open state obtained by addition of the allosteric modulator LY451646 to a suspension of quisqualate-bound, desensitized GluA2. The purpose of the experiment was to test whether structural changes resulting from quisqualate binding to generate the desensitized state could be reversed by addition of an excess of the allosteric modulator LY451646, used to stabilize the open state. The map display shown at left is color-coded to highlight variation in resolution across different regions of the map. b, Density map for the glutamate bound open state obtained by addition of LY451646 30 min prior to agonist, as shown in Figure 2. The map display shown at left is color-coded as in (a) to highlight variation in resolution across different regions of the map. Comparison of the two maps and the fits of ATD and LBD dimers shows that they are essentially identical, establishing the conformational changes that occur with desensitization are reversible and can be modulated by allosteric modulators.
Mentions: To determine whether conformational variability observed for quisqualate-GluA2em complexes reflects the properties of functional receptors, we tested whether subsequent addition of 0.5 mM LY451646 to the same preparation used for cryo-EM analysis of the desensitized state would restore a homogeneous active state conformation. Structural analysis at ~ 16 Å resolution of GluA2em receptor suspensions treated this way demonstrates that the active conformation is indeed restored (Fig. 3c and Extended Data Fig. 6). Our experiments thus establish that in the desensitized state, quisqualate-bound GluA2 is fully functional and capable of undergoing conversion to the active state.

Bottom Line: Desensitization is accompanied by disruption of the amino-terminal domain tetramer in AMPA, but not kainate, receptors with a two-fold to four-fold symmetry transition in the ligand-binding domains in both subtypes.The 7.6 Å structure of a desensitized kainate receptor shows how these changes accommodate channel closing.These findings integrate previous physiological, biochemical and structural analyses of glutamate receptors and provide a molecular explanation for key steps in receptor gating.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Cell Biology, Center for Cancer Research, NCI, NIH, Bethesda, Maryland 20892, USA.

ABSTRACT
Ionotropic glutamate receptors are ligand-gated ion channels that mediate excitatory synaptic transmission in the vertebrate brain. To gain a better understanding of how structural changes gate ion flux across the membrane, we trapped rat AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid) and kainate receptor subtypes in their major functional states and analysed the resulting structures using cryo-electron microscopy. We show that transition to the active state involves a 'corkscrew' motion of the receptor assembly, driven by closure of the ligand-binding domain. Desensitization is accompanied by disruption of the amino-terminal domain tetramer in AMPA, but not kainate, receptors with a two-fold to four-fold symmetry transition in the ligand-binding domains in both subtypes. The 7.6 Å structure of a desensitized kainate receptor shows how these changes accommodate channel closing. These findings integrate previous physiological, biochemical and structural analyses of glutamate receptors and provide a molecular explanation for key steps in receptor gating.

Show MeSH
Related in: MedlinePlus