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- PDB-8ffm: Wildtype rat TRPV2 in nanodiscs bound to RR and 2-APB -

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Basic information

Entry
Database: PDB / ID: 8ffm
TitleWildtype rat TRPV2 in nanodiscs bound to RR and 2-APB
ComponentsTransient receptor potential cation channel subfamily V member 2
KeywordsMEMBRANE PROTEIN / TRPV2 / 2-APB / TRP Channel / Ruthenium Red
Function / homology
Function and homology information


growth cone membrane / TRP channels / response to temperature stimulus / positive regulation of calcium ion import / calcium ion import across plasma membrane / axonal growth cone / positive regulation of axon extension / monoatomic cation channel activity / endomembrane system / calcium channel activity ...growth cone membrane / TRP channels / response to temperature stimulus / positive regulation of calcium ion import / calcium ion import across plasma membrane / axonal growth cone / positive regulation of axon extension / monoatomic cation channel activity / endomembrane system / calcium channel activity / melanosome / lamellipodium / positive regulation of cold-induced thermogenesis / cell body / axon / negative regulation of cell population proliferation / cell surface / identical protein binding / plasma membrane
Similarity search - Function
Transient receptor potential cation channel subfamily V member 1-4 / Transient receptor potential cation channel subfamily V / Ankyrin repeats (3 copies) / Ankyrin repeat profile. / Ankyrin repeat region circular profile. / ankyrin repeats / Ankyrin repeat / Ankyrin repeat-containing domain superfamily / Ion transport domain / Ion transport protein
Similarity search - Domain/homology
2-aminoethyl diphenylborinate / 1,2-DIDECANOYL-SN-GLYCERO-3-PHOSPHOETHANOLAMINE / Chem-R2R / Transient receptor potential cation channel subfamily V member 2
Similarity search - Component
Biological speciesRattus norvegicus (Norway rat)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsPumroy, R.A. / Protopopova, A.D. / Rocereta, J.A. / De Jesus-Perez, J.J. / Fluck, E.C. / Moiseenkova-Bell, V.Y.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM144120 United States
CitationJournal: EMBO Rep / Year: 2024
Title: Molecular details of ruthenium red pore block in TRPV channels.
Authors: Ruth A Pumroy / José J De Jesús-Pérez / Anna D Protopopova / Julia A Rocereta / Edwin C Fluck / Tabea Fricke / Bo-Hyun Lee / Tibor Rohacs / Andreas Leffler / Vera Moiseenkova-Bell /
Abstract: Transient receptor potential vanilloid (TRPV) channels play a critical role in calcium homeostasis, pain sensation, immunological response, and cancer progression. TRPV channels are blocked by ...Transient receptor potential vanilloid (TRPV) channels play a critical role in calcium homeostasis, pain sensation, immunological response, and cancer progression. TRPV channels are blocked by ruthenium red (RR), a universal pore blocker for a wide array of cation channels. Here we use cryo-electron microscopy to reveal the molecular details of RR block in TRPV2 and TRPV5, members of the two TRPV subfamilies. In TRPV2 activated by 2-aminoethoxydiphenyl borate, RR is tightly coordinated in the open selectivity filter, blocking ion flow and preventing channel inactivation. In TRPV5 activated by phosphatidylinositol 4,5-bisphosphate, RR blocks the selectivity filter and closes the lower gate through an interaction with polar residues in the pore vestibule. Together, our results provide a detailed understanding of TRPV subfamily pore block, the dynamic nature of the selectivity filter and allosteric communication between the selectivity filter and lower gate.
History
DepositionDec 9, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Feb 7, 2024Provider: repository / Type: Initial release
Revision 1.1Feb 21, 2024Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Transient receptor potential cation channel subfamily V member 2
B: Transient receptor potential cation channel subfamily V member 2
C: Transient receptor potential cation channel subfamily V member 2
D: Transient receptor potential cation channel subfamily V member 2
hetero molecules


Theoretical massNumber of molelcules
Total (without water)350,74613
Polymers347,1964
Non-polymers3,5509
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Transient receptor potential cation channel subfamily V member 2 / TrpV2 / Osm-9-like TRP channel 2 / OTRPC2 / Stretch-activated channel 2B / Vanilloid receptor-like ...TrpV2 / Osm-9-like TRP channel 2 / OTRPC2 / Stretch-activated channel 2B / Vanilloid receptor-like protein 1 / VRL-1


Mass: 86798.891 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Rattus norvegicus (Norway rat) / Gene: Trpv2, Sac2b, Vrl1 / Production host: Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q9WUD2
#2: Chemical
ChemComp-PEX / 1,2-DIDECANOYL-SN-GLYCERO-3-PHOSPHOETHANOLAMINE


Mass: 522.632 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C25H49NO8P
#3: Chemical
ChemComp-FZ4 / 2-aminoethyl diphenylborinate


Mass: 225.094 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C14H16BNO
#4: Chemical ChemComp-R2R / ruthenium(6+) azanide pentaamino(oxido)ruthenium (1/4/2)


Mass: 559.525 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: H28N14O2Ru3 / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY

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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: Tetramer of wildtype rat TRPV2 in nanodiscs bound to ruthenium red
Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightValue: 0.34 MDa / Experimental value: NO
Source (natural)Organism: Rattus norvegicus (Norway rat)
Source (recombinant)Organism: Saccharomyces cerevisiae (brewer's yeast)
Buffer solutionpH: 8
SpecimenConc.: 2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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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 defocus max: 20000 nm / Nominal defocus min: 5000 nm
Image recordingElectron dose: 42 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 2 / Num. of real images: 10643

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.20.1_4487refinement
PHENIX1.20.1_4487refinement
EM software
IDNameVersionCategory
1Topazparticle selection
4CTFFINDCTF correction
9RELION4initial Euler assignment
10cryoSPARCfinal Euler assignment
13ISOLDEmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 690077
SymmetryPoint symmetry: C4 (4 fold cyclic)
3D reconstructionResolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 65741 / Symmetry type: POINT
Atomic model buildingPDB-ID: 7n0m

7n0m


Accession code: 7n0m / Source name: PDB / Type: experimental model
RefinementCross valid method: NONE

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