Limits...
NMDA receptor structures reveal subunit arrangement and pore architecture.

Lee CH, Lü W, Michel JC, Goehring A, Du J, Song X, Gouaux E - Nature (2014)

Bottom Line: Receptor subunits are arranged in a 1-2-1-2 fashion, demonstrating extensive interactions between the amino-terminal and ligand-binding domains.The transmembrane domains harbour a closed-blocked ion channel, a pyramidal central vestibule lined by residues implicated in binding ion channel blockers and magnesium, and a ∼twofold symmetric arrangement of ion channel pore loops.These structures provide new insights into the architecture, allosteric coupling and ion channel function of NMDA receptors.

View Article: PubMed Central - PubMed

Affiliation: 1] Vollum Institute, Oregon Health & Science University, 3181 SW Sam Jackson Park Road, Portland, Oregon 97239, USA [2].

ABSTRACT
N-methyl-d-aspartate (NMDA) receptors are Hebbian-like coincidence detectors, requiring binding of glycine and glutamate in combination with the relief of voltage-dependent magnesium block to open an ion conductive pore across the membrane bilayer. Despite the importance of the NMDA receptor in the development and function of the brain, a molecular structure of an intact receptor has remained elusive. Here we present X-ray crystal structures of the Xenopus laevis GluN1-GluN2B NMDA receptor with the allosteric inhibitor, Ro25-6981, partial agonists and the ion channel blocker, MK-801. Receptor subunits are arranged in a 1-2-1-2 fashion, demonstrating extensive interactions between the amino-terminal and ligand-binding domains. The transmembrane domains harbour a closed-blocked ion channel, a pyramidal central vestibule lined by residues implicated in binding ion channel blockers and magnesium, and a ∼twofold symmetric arrangement of ion channel pore loops. These structures provide new insights into the architecture, allosteric coupling and ion channel function of NMDA receptors.

Show MeSH

Related in: MedlinePlus

Transmembrane domain architecture, symmetry and coupling to LBDa, View of the TMD parallel to the membrane. GluN1 subunits are blue and the GluN2B subunits are orange. b, View of the TMD, along the pore axis, from the cytoplasmic side of the membrane. c, View of a solvent accessible surface carved along the pore axis using the computer program HOLE, parallel to the membrane, showing that the M3 bundle crossing near the extracellular side of the membrane and the entry into the selectivity filter region, from the central aqueous vestibule, form constrictions in the pore. The color coding for the dots that indicate the pore radius is 1.15 Å < green < 2.3 Å < blue. Because a number of side chains are not included in the structure, due to the moderate resolution of the diffraction data, the size of the pore is approximate. d, View of the extracellular ends of the M3 helices of the NMDA receptor. We have highlighted as spheres the α-carbon atoms for residues Thr 646 and Ala 645 in the GluN1/GluN2B structure, respectively. The distances between neighboring atoms are 6.2, 8.0, 5.4 and 7.1 Å, starting from the α-carbon of GluN2B on the left and going clockwise. e, View of the intracellular ends of the TMD of the NMDA receptor in comparison with KcsA. Here, the M2 helices of the NMDA receptor were superimposed on the corresponding helices in KcsA, showing the deviation from 4-fold symmetry. f, Side view of the TMD showing a positive electron density feature (green mesh) in the central vestibule, calculated using Fo-Fc coefficients and phases from the refined structure. The map is contoured at 2.8 σ. Data set 2 and Structure 2 were employed in all panels (Extended Data Table 2).
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC4263351&req=5

Figure 5: Transmembrane domain architecture, symmetry and coupling to LBDa, View of the TMD parallel to the membrane. GluN1 subunits are blue and the GluN2B subunits are orange. b, View of the TMD, along the pore axis, from the cytoplasmic side of the membrane. c, View of a solvent accessible surface carved along the pore axis using the computer program HOLE, parallel to the membrane, showing that the M3 bundle crossing near the extracellular side of the membrane and the entry into the selectivity filter region, from the central aqueous vestibule, form constrictions in the pore. The color coding for the dots that indicate the pore radius is 1.15 Å < green < 2.3 Å < blue. Because a number of side chains are not included in the structure, due to the moderate resolution of the diffraction data, the size of the pore is approximate. d, View of the extracellular ends of the M3 helices of the NMDA receptor. We have highlighted as spheres the α-carbon atoms for residues Thr 646 and Ala 645 in the GluN1/GluN2B structure, respectively. The distances between neighboring atoms are 6.2, 8.0, 5.4 and 7.1 Å, starting from the α-carbon of GluN2B on the left and going clockwise. e, View of the intracellular ends of the TMD of the NMDA receptor in comparison with KcsA. Here, the M2 helices of the NMDA receptor were superimposed on the corresponding helices in KcsA, showing the deviation from 4-fold symmetry. f, Side view of the TMD showing a positive electron density feature (green mesh) in the central vestibule, calculated using Fo-Fc coefficients and phases from the refined structure. The map is contoured at 2.8 σ. Data set 2 and Structure 2 were employed in all panels (Extended Data Table 2).

Mentions: The electron density associated with Data set 2/Structure 2 allowed us to position the polypeptide main chain for the M1-M4 helices of all subunits (Supplementary Video 1). To trace the polypeptide associated with the pore loop, we exploited the continuous electron density for this region in the GluN2B subunit D and, by applying non crystallographic symmetry defined by the transmembrane segments of the other subunits, we traced the three remaining pore loops (Fig. 5a, 5b). The arrangement of transmembrane helices is like that of the GluA2 AMPA receptor28 (Extended Data Fig. 7a), although in the NMDA receptor we have a more complete representation of the ion channel pore and putative selectivity filter. The pre-M1 region of the NMDA receptor forms a ‘collar’ around the extracellular regions of the M3 helices, residing near the boundary of the extracellular side of the membrane. The M1 helix descends across the membrane and makes interactions with the pore-lining M3 helix of the same subunit and the M4 helix of a neighbor. Electron density for the cytoplasmic loop connecting M1 to M2 is weak or missing, and thus this region is absent from the structure. We can visualize the M2 pore helix and most of the extended region of the pore loop forming the selectivity filter and its connection to the N-terminus of M3.


NMDA receptor structures reveal subunit arrangement and pore architecture.

Lee CH, Lü W, Michel JC, Goehring A, Du J, Song X, Gouaux E - Nature (2014)

Transmembrane domain architecture, symmetry and coupling to LBDa, View of the TMD parallel to the membrane. GluN1 subunits are blue and the GluN2B subunits are orange. b, View of the TMD, along the pore axis, from the cytoplasmic side of the membrane. c, View of a solvent accessible surface carved along the pore axis using the computer program HOLE, parallel to the membrane, showing that the M3 bundle crossing near the extracellular side of the membrane and the entry into the selectivity filter region, from the central aqueous vestibule, form constrictions in the pore. The color coding for the dots that indicate the pore radius is 1.15 Å < green < 2.3 Å < blue. Because a number of side chains are not included in the structure, due to the moderate resolution of the diffraction data, the size of the pore is approximate. d, View of the extracellular ends of the M3 helices of the NMDA receptor. We have highlighted as spheres the α-carbon atoms for residues Thr 646 and Ala 645 in the GluN1/GluN2B structure, respectively. The distances between neighboring atoms are 6.2, 8.0, 5.4 and 7.1 Å, starting from the α-carbon of GluN2B on the left and going clockwise. e, View of the intracellular ends of the TMD of the NMDA receptor in comparison with KcsA. Here, the M2 helices of the NMDA receptor were superimposed on the corresponding helices in KcsA, showing the deviation from 4-fold symmetry. f, Side view of the TMD showing a positive electron density feature (green mesh) in the central vestibule, calculated using Fo-Fc coefficients and phases from the refined structure. The map is contoured at 2.8 σ. Data set 2 and Structure 2 were employed in all panels (Extended Data Table 2).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4263351&req=5

Figure 5: Transmembrane domain architecture, symmetry and coupling to LBDa, View of the TMD parallel to the membrane. GluN1 subunits are blue and the GluN2B subunits are orange. b, View of the TMD, along the pore axis, from the cytoplasmic side of the membrane. c, View of a solvent accessible surface carved along the pore axis using the computer program HOLE, parallel to the membrane, showing that the M3 bundle crossing near the extracellular side of the membrane and the entry into the selectivity filter region, from the central aqueous vestibule, form constrictions in the pore. The color coding for the dots that indicate the pore radius is 1.15 Å < green < 2.3 Å < blue. Because a number of side chains are not included in the structure, due to the moderate resolution of the diffraction data, the size of the pore is approximate. d, View of the extracellular ends of the M3 helices of the NMDA receptor. We have highlighted as spheres the α-carbon atoms for residues Thr 646 and Ala 645 in the GluN1/GluN2B structure, respectively. The distances between neighboring atoms are 6.2, 8.0, 5.4 and 7.1 Å, starting from the α-carbon of GluN2B on the left and going clockwise. e, View of the intracellular ends of the TMD of the NMDA receptor in comparison with KcsA. Here, the M2 helices of the NMDA receptor were superimposed on the corresponding helices in KcsA, showing the deviation from 4-fold symmetry. f, Side view of the TMD showing a positive electron density feature (green mesh) in the central vestibule, calculated using Fo-Fc coefficients and phases from the refined structure. The map is contoured at 2.8 σ. Data set 2 and Structure 2 were employed in all panels (Extended Data Table 2).
Mentions: The electron density associated with Data set 2/Structure 2 allowed us to position the polypeptide main chain for the M1-M4 helices of all subunits (Supplementary Video 1). To trace the polypeptide associated with the pore loop, we exploited the continuous electron density for this region in the GluN2B subunit D and, by applying non crystallographic symmetry defined by the transmembrane segments of the other subunits, we traced the three remaining pore loops (Fig. 5a, 5b). The arrangement of transmembrane helices is like that of the GluA2 AMPA receptor28 (Extended Data Fig. 7a), although in the NMDA receptor we have a more complete representation of the ion channel pore and putative selectivity filter. The pre-M1 region of the NMDA receptor forms a ‘collar’ around the extracellular regions of the M3 helices, residing near the boundary of the extracellular side of the membrane. The M1 helix descends across the membrane and makes interactions with the pore-lining M3 helix of the same subunit and the M4 helix of a neighbor. Electron density for the cytoplasmic loop connecting M1 to M2 is weak or missing, and thus this region is absent from the structure. We can visualize the M2 pore helix and most of the extended region of the pore loop forming the selectivity filter and its connection to the N-terminus of M3.

Bottom Line: Receptor subunits are arranged in a 1-2-1-2 fashion, demonstrating extensive interactions between the amino-terminal and ligand-binding domains.The transmembrane domains harbour a closed-blocked ion channel, a pyramidal central vestibule lined by residues implicated in binding ion channel blockers and magnesium, and a ∼twofold symmetric arrangement of ion channel pore loops.These structures provide new insights into the architecture, allosteric coupling and ion channel function of NMDA receptors.

View Article: PubMed Central - PubMed

Affiliation: 1] Vollum Institute, Oregon Health & Science University, 3181 SW Sam Jackson Park Road, Portland, Oregon 97239, USA [2].

ABSTRACT
N-methyl-d-aspartate (NMDA) receptors are Hebbian-like coincidence detectors, requiring binding of glycine and glutamate in combination with the relief of voltage-dependent magnesium block to open an ion conductive pore across the membrane bilayer. Despite the importance of the NMDA receptor in the development and function of the brain, a molecular structure of an intact receptor has remained elusive. Here we present X-ray crystal structures of the Xenopus laevis GluN1-GluN2B NMDA receptor with the allosteric inhibitor, Ro25-6981, partial agonists and the ion channel blocker, MK-801. Receptor subunits are arranged in a 1-2-1-2 fashion, demonstrating extensive interactions between the amino-terminal and ligand-binding domains. The transmembrane domains harbour a closed-blocked ion channel, a pyramidal central vestibule lined by residues implicated in binding ion channel blockers and magnesium, and a ∼twofold symmetric arrangement of ion channel pore loops. These structures provide new insights into the architecture, allosteric coupling and ion channel function of NMDA receptors.

Show MeSH
Related in: MedlinePlus