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- PDB-9c2c: Cryo-EM structure of the human P2X1 receptor in the NF449-bound i... -

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Basic information

Entry
Database: PDB / ID: 9c2c
TitleCryo-EM structure of the human P2X1 receptor in the NF449-bound inhibited state
ComponentsP2X purinoceptor 1
KeywordsMEMBRANE PROTEIN / Ion Channel / Ligand-gated Ion Channel / P2X Receptor / Allosteric Antagonist
Function / homology
Function and homology information


regulation of vascular associated smooth muscle contraction / Platelet homeostasis / insemination / positive regulation of calcium ion import across plasma membrane / purinergic nucleotide receptor activity / extracellularly ATP-gated monoatomic cation channel activity / suramin binding / serotonin secretion by platelet / Elevation of cytosolic Ca2+ levels / regulation of presynaptic cytosolic calcium ion concentration ...regulation of vascular associated smooth muscle contraction / Platelet homeostasis / insemination / positive regulation of calcium ion import across plasma membrane / purinergic nucleotide receptor activity / extracellularly ATP-gated monoatomic cation channel activity / suramin binding / serotonin secretion by platelet / Elevation of cytosolic Ca2+ levels / regulation of presynaptic cytosolic calcium ion concentration / ligand-gated calcium channel activity / ceramide biosynthetic process / regulation of synaptic vesicle exocytosis / response to ATP / neuronal action potential / specific granule membrane / monoatomic cation channel activity / monoatomic ion transport / presynaptic active zone membrane / secretory granule membrane / synaptic transmission, glutamatergic / calcium ion transmembrane transport / platelet activation / regulation of blood pressure / postsynaptic membrane / membrane raft / external side of plasma membrane / glutamatergic synapse / Neutrophil degranulation / protein-containing complex binding / apoptotic process / signal transduction / protein-containing complex / ATP binding / identical protein binding / plasma membrane
Similarity search - Function
P2X1 purinoceptor / : / ATP P2X receptor / ATP P2X receptors signature. / P2X purinoreceptor / P2X purinoreceptor extracellular domain superfamily
Similarity search - Domain/homology
: / P2X purinoceptor 1
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsOken, A.C. / Lisi, N.E. / Ditter, I.A. / Shi, H. / Mansoor, S.E.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)R00HL138129 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)DP2GM149551 United States
CitationJournal: Nat Commun / Year: 2024
Title: Cryo-EM structures of the human P2X1 receptor reveal subtype-specific architecture and antagonism by supramolecular ligand-binding.
Authors: Adam C Oken / Nicolas E Lisi / Ismayn A Ditter / Haoyuan Shi / Nadia A Nechiporuk / Steven E Mansoor /
Abstract: P2X receptors are a family of seven trimeric non-selective cation channels that are activated by extracellular ATP to play roles in the cardiovascular, neuronal, and immune systems. Although it is ...P2X receptors are a family of seven trimeric non-selective cation channels that are activated by extracellular ATP to play roles in the cardiovascular, neuronal, and immune systems. Although it is known that the P2X1 receptor subtype has increased sensitivity to ATP and fast desensitization kinetics, an underlying molecular explanation for these subtype-selective features is lacking. Here we report high-resolution cryo-EM structures of the human P2X1 receptor in the apo closed, ATP-bound desensitized, and the high-affinity antagonist NF449-bound inhibited states. The apo closed and ATP-bound desensitized state structures of human P2X1 define subtype-specific properties such as distinct pore architecture and ATP-interacting residues. The NF449-bound inhibited state structure of human P2X1 reveals that NF449 has a unique dual-ligand supramolecular binding mode at the interface of neighboring protomers, inhibiting channel activation by overlapping with the canonical P2X receptor ATP-binding site. Altogether, these data define the molecular pharmacology of the human P2X1 receptor laying the foundation for structure-based drug design.
History
DepositionMay 30, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 9, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: P2X purinoceptor 1
C: P2X purinoceptor 1
B: P2X purinoceptor 1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)145,08318
Polymers135,1173
Non-polymers9,96615
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein P2X purinoceptor 1 / P2X1 / ATP receptor / Purinergic receptor


Mass: 45038.957 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: P2RX1, P2X1 / Cell (production host): HEK293 GNTI- / Production host: Homo sapiens (human) / References: UniProt: P51575
#2: Sugar
ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 9 / Source method: obtained synthetically / Formula: C8H15NO6
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0
#3: Chemical
ChemComp-A1ALI / 4,4',4'',4'''-{carbonylbis[azanediylbenzene-5,1,3-triylbis(carbonylazanediyl)]}tetra(benzene-1,3-disulfonic acid)


Mass: 1329.236 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C41H32N6O29S8 / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY
Has protein modificationY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Membrane protein / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human) / Cell: HEK293 GNTI-
Buffer solutionpH: 7
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 130000 X / Nominal defocus max: 1400 nm / Nominal defocus min: 900 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm
Specimen holderSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 45 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of real images: 22866
EM imaging opticsEnergyfilter slit width: 20 eV

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C3 (3 fold cyclic)
3D reconstructionResolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 86114 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0067863
ELECTRON MICROSCOPYf_angle_d0.83110776
ELECTRON MICROSCOPYf_dihedral_angle_d9.651083
ELECTRON MICROSCOPYf_chiral_restr0.0521125
ELECTRON MICROSCOPYf_plane_restr0.0051314

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