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- PDB-3j5q: Structure of TRPV1 ion channel in complex with DkTx and RTX deter... -

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Basic information

Entry
Database: PDB / ID: 3j5q
TitleStructure of TRPV1 ion channel in complex with DkTx and RTX determined by single particle electron cryo-microscopy
Components
  • Kappa-theraphotoxin-Cg1a 1
  • Transient receptor potential cation channel subfamily V member 1TRPV1
KeywordsTRANSPORT PROTEIN/TOXIN / TRPV1 channel / DkTx / RTX / TRANSPORT PROTEIN-TOXIN complex
Function / homology
Function and homology information


temperature-gated ion channel activity / response to capsazepine / excitatory extracellular ligand-gated monoatomic ion channel activity / negative regulation of establishment of blood-brain barrier / sensory perception of mechanical stimulus / peptide secretion / urinary bladder smooth muscle contraction / detection of chemical stimulus involved in sensory perception of pain / cellular response to temperature stimulus / smooth muscle contraction involved in micturition ...temperature-gated ion channel activity / response to capsazepine / excitatory extracellular ligand-gated monoatomic ion channel activity / negative regulation of establishment of blood-brain barrier / sensory perception of mechanical stimulus / peptide secretion / urinary bladder smooth muscle contraction / detection of chemical stimulus involved in sensory perception of pain / cellular response to temperature stimulus / smooth muscle contraction involved in micturition / TRP channels / cellular response to acidic pH / thermoception / fever generation / detection of temperature stimulus involved in thermoception / dendritic spine membrane / glutamate secretion / negative regulation of systemic arterial blood pressure / chloride channel regulator activity / response to pH / monoatomic cation transmembrane transporter activity / cellular response to ATP / response to pain / temperature homeostasis / ion channel inhibitor activity / negative regulation of heart rate / detection of temperature stimulus involved in sensory perception of pain / cellular response to alkaloid / diet induced thermogenesis / extracellular ligand-gated monoatomic ion channel activity / behavioral response to pain / intracellularly gated calcium channel activity / cellular response to cytokine stimulus / negative regulation of mitochondrial membrane potential / calcium ion import across plasma membrane / ligand-gated monoatomic ion channel activity / sodium channel regulator activity / potassium channel regulator activity / monoatomic cation channel activity / sensory perception of pain / response to organonitrogen compound / monoatomic ion transmembrane transport / phosphatidylinositol binding / cellular response to nerve growth factor stimulus / calcium ion transmembrane transport / microglial cell activation / calcium channel activity / lipid metabolic process / response to peptide hormone / phosphoprotein binding / cellular response to growth factor stimulus / calcium ion transport / positive regulation of nitric oxide biosynthetic process / transmembrane signaling receptor activity / cellular response to heat / cellular response to tumor necrosis factor / positive regulation of cytosolic calcium ion concentration / response to heat / toxin activity / postsynaptic membrane / protein homotetramerization / calmodulin binding / neuron projection / positive regulation of apoptotic process / external side of plasma membrane / neuronal cell body / dendrite / negative regulation of transcription by RNA polymerase II / extracellular region / ATP binding / membrane / metal ion binding / identical protein binding / plasma membrane
Similarity search - Function
Huwentoxin-1 family / Ion channel inhibitory toxin / Transient receptor potential cation channel subfamily V member 1-4 / Ankyrin repeat-containing domain / Transient receptor potential cation channel subfamily V / Ankyrin repeat / Ankyrin repeats (3 copies) / Ankyrin repeat profile. / Ankyrin repeat region circular profile. / ankyrin repeats ...Huwentoxin-1 family / Ion channel inhibitory toxin / Transient receptor potential cation channel subfamily V member 1-4 / Ankyrin repeat-containing domain / Transient receptor potential cation channel subfamily V / Ankyrin repeat / Ankyrin repeats (3 copies) / Ankyrin repeat profile. / Ankyrin repeat region circular profile. / ankyrin repeats / Ankyrin repeat / Ankyrin repeat-containing domain superfamily / Serine Threonine Protein Phosphatase 5, Tetratricopeptide repeat / Ion transport domain / Ion transport protein / Alpha Horseshoe / Mainly Alpha
Similarity search - Domain/homology
Transient receptor potential cation channel subfamily V member 1 / Kappa-theraphotoxin-Cg1a 1
Similarity search - Component
Biological speciesRattus norvegicus (Norway rat)
Chilobrachys guangxiensis (spider)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsLiao, M. / Cao, E. / Julius, D. / Cheng, Y.
CitationJournal: Nature / Year: 2013
Title: TRPV1 structures in distinct conformations reveal activation mechanisms.
Authors: Erhu Cao / Maofu Liao / Yifan Cheng / David Julius /
Abstract: Transient receptor potential (TRP) channels are polymodal signal detectors that respond to a wide range of physical and chemical stimuli. Elucidating how these channels integrate and convert ...Transient receptor potential (TRP) channels are polymodal signal detectors that respond to a wide range of physical and chemical stimuli. Elucidating how these channels integrate and convert physiological signals into channel opening is essential to understanding how they regulate cell excitability under normal and pathophysiological conditions. Here we exploit pharmacological probes (a peptide toxin and small vanilloid agonists) to determine structures of two activated states of the capsaicin receptor, TRPV1. A domain (consisting of transmembrane segments 1-4) that moves during activation of voltage-gated channels remains stationary in TRPV1, highlighting differences in gating mechanisms for these structurally related channel superfamilies. TRPV1 opening is associated with major structural rearrangements in the outer pore, including the pore helix and selectivity filter, as well as pronounced dilation of a hydrophobic constriction at the lower gate, suggesting a dual gating mechanism. Allosteric coupling between upper and lower gates may account for rich physiological modulation exhibited by TRPV1 and other TRP channels.
History
DepositionOct 28, 2013Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 4, 2013Provider: repository / Type: Initial release
Revision 1.1Dec 11, 2013Group: Database references
Revision 1.2Dec 18, 2013Group: Database references
Revision 1.3Jul 18, 2018Group: Data collection / Category: em_software / Item: _em_software.image_processing_id / _em_software.name
Revision 2.0Nov 2, 2022Group: Advisory / Atomic model ...Advisory / Atomic model / Data collection / Database references / Derived calculations / Polymer sequence / Source and taxonomy / Structure summary
Category: atom_site / database_2 ...atom_site / database_2 / entity / entity_name_com / entity_poly / entity_poly_seq / entity_src_gen / entity_src_nat / pdbx_entry_details / pdbx_poly_seq_scheme / pdbx_struct_sheet_hbond / pdbx_unobs_or_zero_occ_atoms / pdbx_unobs_or_zero_occ_residues / pdbx_validate_close_contact / pdbx_validate_torsion / struct_mon_prot_cis / struct_ref / struct_ref_seq / struct_ref_seq_dif / struct_sheet / struct_sheet_order / struct_sheet_range
Item: _atom_site.Cartn_x / _atom_site.Cartn_y ..._atom_site.Cartn_x / _atom_site.Cartn_y / _atom_site.Cartn_z / _atom_site.auth_atom_id / _atom_site.auth_comp_id / _atom_site.label_atom_id / _atom_site.label_comp_id / _atom_site.label_seq_id / _atom_site.type_symbol / _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _entity.formula_weight / _entity.pdbx_description / _entity.pdbx_fragment / _entity_name_com.name / _entity_poly.pdbx_seq_one_letter_code / _entity_poly.pdbx_seq_one_letter_code_can / _entity_src_nat.common_name / _entity_src_nat.pdbx_beg_seq_num / _entity_src_nat.pdbx_end_seq_num / _entity_src_nat.pdbx_organism_scientific / _pdbx_entry_details.sequence_details / _pdbx_validate_close_contact.auth_comp_id_1 / _pdbx_validate_close_contact.auth_comp_id_2 / _pdbx_validate_torsion.auth_comp_id / _struct_mon_prot_cis.auth_comp_id / _struct_mon_prot_cis.label_comp_id / _struct_mon_prot_cis.label_seq_id / _struct_mon_prot_cis.pdbx_auth_comp_id_2 / _struct_mon_prot_cis.pdbx_label_comp_id_2 / _struct_mon_prot_cis.pdbx_label_seq_id_2

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Structure visualization

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Assembly

Deposited unit
D: Transient receptor potential cation channel subfamily V member 1
B: Transient receptor potential cation channel subfamily V member 1
E: Transient receptor potential cation channel subfamily V member 1
G: Transient receptor potential cation channel subfamily V member 1
A: Kappa-theraphotoxin-Cg1a 1
C: Kappa-theraphotoxin-Cg1a 1
F: Kappa-theraphotoxin-Cg1a 1
H: Kappa-theraphotoxin-Cg1a 1


Theoretical massNumber of molelcules
Total (without water)296,9138
Polymers296,9138
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein
Transient receptor potential cation channel subfamily V member 1 / TRPV1 / TrpV1 / Capsaicin receptor / Osm-9-like TRP channel 1 / OTRPC1 / Vanilloid receptor 1 / Vanilloid ...TrpV1 / Capsaicin receptor / Osm-9-like TRP channel 1 / OTRPC1 / Vanilloid receptor 1 / Vanilloid receptor type 1-like


Mass: 70795.211 Da / Num. of mol.: 4 / Fragment: SEE REMARK 999
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Rattus norvegicus (Norway rat) / Gene: Trpv1, Vr1, Vr1l / Plasmid: pFastBac1 / Cell line (production host): HEK293S GnTI / Production host: Homo sapiens (human) / References: UniProt: O35433
#2: Protein/peptide
Kappa-theraphotoxin-Cg1a 1 / double-knot toxin


Mass: 3433.014 Da / Num. of mol.: 4 / Fragment: UNP residues 51-81 / Source method: isolated from a natural source / Source: (natural) Chilobrachys guangxiensis (spider) / References: UniProt: P0C247
Sequence detailsTHE TRPV1 CONSTRUCT COMPRISES RESIDUES 110-603 AND 627-764, WITH RESIDUES 604-626 DELETED. RESIDUES ...THE TRPV1 CONSTRUCT COMPRISES RESIDUES 110-603 AND 627-764, WITH RESIDUES 604-626 DELETED. RESIDUES 719-764 ARE NOT MODELED, WITH THE EXCEPTION OF 11 RESIDUES (NUMBERED 752-762 IN THE COORDINATES) WHOSE IDENTITIES ARE NOT KNOWN.

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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: Rat TRPV1 in complex with DkTx and resiniferatoxin / Type: COMPLEX / Details: Tetramer
Molecular weightValue: 0.3 MDa / Experimental value: NO
Buffer solutionName: 150 mM NaCl, 20 mM HEPES, 2 mM TCEP / pH: 7.4 / Details: 150 mM NaCl, 20 mM HEPES, 2 mM TCEP
SpecimenConc.: 0.3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: 400 mesh Quantifoil grid
VitrificationInstrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Temp: 120 K / Humidity: 90 %
Details: Blot for 6 seconds before plunging into liquid ethane (FEI VITROBOT MARK III)
Method: Blot for 6 sec

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Electron microscopy imaging

Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company
MicroscopyModel: FEI POLARA 300 / Date: Jan 1, 2013
Details: Gatan K2 Summit operated in super-resolution counting mode, image recorded with dose fractionation method.
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 31000 X / Calibrated magnification: 31000 X / Nominal defocus max: 3000 nm / Nominal defocus min: 1500 nm / Cs: 2 mm / Camera length: 0 mm
Specimen holderSpecimen holder model: OTHER / Specimen holder type: FEI Polara cartridge
Image recordingElectron dose: 21 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)
Details: Operated in super-resolution counting mode, dose fractionation
Image scansNum. digital images: 900
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthRelative weight: 1

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Processing

EM softwareName: RELION / Category: 3D reconstruction
CTF correctionDetails: Each particle
SymmetryPoint symmetry: C4 (4 fold cyclic)
3D reconstructionMethod: Maximum likelihood / Resolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 36158 / Nominal pixel size: 1.2156 Å / Actual pixel size: 1.2156 Å
Details: The entire ankyrin structure was fitted as a single rigid body. As a result, the first two ankyrin repeats (UNP residues 111-199) are included but are not well defined by the experimental ...Details: The entire ankyrin structure was fitted as a single rigid body. As a result, the first two ankyrin repeats (UNP residues 111-199) are included but are not well defined by the experimental density. Interatomic clashes between the ankyrin repeats and the C-terminal beta sheet are the result of poor geometry due to relatively poor experimental density in these regions. Resolution and averaging of DkTx density was insufficient to provide side chain geometry. The DkTx model was generated as a poly-ALA model based on hanatoxin (PDB entry 1D1H) and docked into the DkTx density with slight modification. (Single particle details: 3D classification, refinement, and reconstruction were performed using RELION) (Single particle--Applied symmetry: C4)
Refinement type: HALF-MAPS REFINED INDEPENDENTLY / Symmetry type: POINT
Atomic model buildingProtocol: AB INITIO MODEL / Space: REAL
Details: REFINEMENT PROTOCOL--de novo model building DETAILS--The entire ankyrin structure was fitted as a single rigid body. As a result, the first two ankyrin repeats (UNP residues 111-199) are ...Details: REFINEMENT PROTOCOL--de novo model building DETAILS--The entire ankyrin structure was fitted as a single rigid body. As a result, the first two ankyrin repeats (UNP residues 111-199) are included but are not well defined by the experimental density.
Refinement stepCycle: LAST
ProteinNucleic acidLigandSolventTotal
Num. atoms18264 0 0 0 18264

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