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- EMDB-36671: Cryo-EM structure of the panda P2X7 receptor in complex with PPNDS -

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Basic information

Entry
Database: EMDB / ID: EMD-36671
TitleCryo-EM structure of the panda P2X7 receptor in complex with PPNDS
Map data
Sample
  • Complex: P2X7 trimer
    • Protein or peptide: P2X purinoceptor
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: 3-[(E)-{4-formyl-5-hydroxy-6-methyl-3-[(phosphonooxy)methyl]pyridin-2-yl}diazenyl]-7-nitronaphthalene-1,5-disulfonic acid
  • Ligand: water
KeywordsChannel / TRANSPORT PROTEIN
Function / homology
Function and homology information


extracellularly ATP-gated monoatomic cation channel activity / purinergic nucleotide receptor activity / response to ATP / calcium ion transmembrane transport / postsynapse / ATP binding / plasma membrane
Similarity search - Function
P2X purinoreceptor 7, intracellular domain / P2X purinoreceptor 7 intracellular domain / P2X7 purinoceptor / : / ATP P2X receptors signature. / ATP P2X receptor / P2X purinoreceptor / P2X purinoreceptor extracellular domain superfamily
Similarity search - Domain/homology
Biological speciesAiluropoda melanoleuca (giant panda)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.34 Å
AuthorsSheng D / Hattori M
Funding support China, 1 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC) China
Citation
Journal: Elife / Year: 2024
Title: Structural insights into the orthosteric inhibition of P2X receptors by non-ATP analog antagonists.
Authors: Danqi Sheng / Chen-Xi Yue / Fei Jin / Yao Wang / Muneyoshi Ichikawa / Ye Yu / Chang-Run Guo / Motoyuki Hattori /
Abstract: P2X receptors are extracellular ATP-gated ion channels that form homo- or heterotrimers and consist of seven subtypes. They are expressed in various tissues, including neuronal and nonneuronal cells, ...P2X receptors are extracellular ATP-gated ion channels that form homo- or heterotrimers and consist of seven subtypes. They are expressed in various tissues, including neuronal and nonneuronal cells, and play critical roles in physiological processes such as neurotransmission, inflammation, pain, and cancer. As a result, P2X receptors have attracted considerable interest as drug targets, and various competitive inhibitors have been developed. However, although several P2X receptor structures from different subtypes have been reported, the limited structural information of P2X receptors in complex with competitive antagonists hampers the understanding of orthosteric inhibition, hindering the further design and optimization of those antagonists for drug discovery. We determined the cryogenic electron microscopy (cryo-EM) structures of the mammalian P2X7 receptor in complex with two classical competitive antagonists of pyridoxal-5'-phosphate derivatives, pyridoxal-5'-phosphate-6-(2'-naphthylazo-6'-nitro-4',8'-disulfonate) (PPNDS) and pyridoxal phosphate-6-azophenyl-2',5'-disulfonic acid (PPADS), and performed structure-based mutational analysis by patch-clamp recording as well as molecular dynamics (MD) simulations. Our structures revealed the orthosteric site for PPADS/PPNDS, and structural comparison with the previously reported apo- and ATP-bound structures showed how PPADS/PPNDS binding inhibits the conformational changes associated with channel activation. In addition, structure-based mutational analysis identified key residues involved in the PPNDS sensitivity of P2X1 and P2X3, which are known to have higher affinity for PPADS/PPNDS than other P2X subtypes.
#1: Journal: Elife / Year: 2023
Title: Structural insights into the orthosteric inhibition of P2X receptors by non-ATP-analog antagonists
Authors: Sheng D / Yue C / Jin F / Wang Y / Ichikawa M / Yu Y / Guo CR / Hattori M
History
DepositionJun 27, 2023-
Header (metadata) releaseNov 29, 2023-
Map releaseNov 29, 2023-
UpdateOct 9, 2024-
Current statusOct 9, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_36671.png

Downloads & links

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Map

FileDownload / File: emd_36671.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 256 pix.
= 212.48 Å		0.83 Å/pix.
x 256 pix.
= 212.48 Å		0.83 Å/pix.
x 256 pix.
= 212.48 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.83 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.25
Minimum - Maximum-0.6571219 - 1.2528806
Average (Standard dev.)0.00069344515 (±0.03781402)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 212.48 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_36671_half_map_1.map
Projections & Slices
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Half map: #2

Fileemd_36671_half_map_2.map
Projections & Slices
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Histogram (log scale)

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Sample components

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Entire : P2X7 trimer

EntireName: P2X7 trimer
Components
  • Complex: P2X7 trimer
    • Protein or peptide: P2X purinoceptor
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: 3-[(E)-{4-formyl-5-hydroxy-6-methyl-3-[(phosphonooxy)methyl]pyridin-2-yl}diazenyl]-7-nitronaphthalene-1,5-disulfonic acid
  • Ligand: water

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Supramolecule #1: P2X7 trimer

SupramoleculeName: P2X7 trimer / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Ailuropoda melanoleuca (giant panda)

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Macromolecule #1: P2X purinoceptor

MacromoleculeName: P2X purinoceptor / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Ailuropoda melanoleuca (giant panda)
Molecular weightTheoretical: 38.716234 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: GSSTNYGTIK WIFHALVFSY ISFALISDKR YQKKEPLISS VHTKVKGIAE VKAEILENGM KKMVSGVFDT ADYTFPLQGN SFFVMTNFI KTEGQQQGLC PDFPTARTIC SSDRGCKKGR MDPQSKGIQT GRCVVYKERL KTCEVSAWCP IEEVKDAPRP A LLNSAENF ...String:
GSSTNYGTIK WIFHALVFSY ISFALISDKR YQKKEPLISS VHTKVKGIAE VKAEILENGM KKMVSGVFDT ADYTFPLQGN SFFVMTNFI KTEGQQQGLC PDFPTARTIC SSDRGCKKGR MDPQSKGIQT GRCVVYKERL KTCEVSAWCP IEEVKDAPRP A LLNSAENF TVLIKNNIDF PGHNYTTRNI LPGVNITCTF HKTQNPQCPI FRLGDIFQET GDSFSDVAIQ GGIMGIEIYW DC NLDGWFH HCRPKYSFRR LDDKTTSESL YPGYNFRYAK YYKENNVEKR TLIKVFGIRF DILVFGTGGK FNVIQLAVYI GSV ISYFGL ATVFIDILIN TYSASS

UniProtKB: P2X purinoceptor

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Macromolecule #2: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 2 / Number of copies: 6 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose

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Macromolecule #3: 3-[(E)-{4-formyl-5-hydroxy-6-methyl-3-[(phosphonooxy)methyl]pyrid...

MacromoleculeName: 3-[(E)-{4-formyl-5-hydroxy-6-methyl-3-[(phosphonooxy)methyl]pyridin-2-yl}diazenyl]-7-nitronaphthalene-1,5-disulfonic acid
type: ligand / ID: 3 / Number of copies: 3 / Formula: 20V
Molecular weightTheoretical: 606.434 Da

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Macromolecule #4: water

MacromoleculeName: water / type: ligand / ID: 4 / Number of copies: 3 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.3 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER / Details: CryoSPARC ab-initio reconstruction
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.34 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 121008
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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