3J5Q

Structure of TRPV1 ion channel in complex with DkTx and RTX determined by single particle electron cryo-microscopy


ELECTRON MICROSCOPY
Sample
Rat TRPV1 in complex with DkTx and resiniferatoxin
Specimen Preparation
Sample Aggregation StatePARTICLE
Vitrification InstrumentFEI VITROBOT MARK III
Cryogen NameETHANE
Sample Vitrification DetailsBlot for 6 seconds before plunging into liquid ethane (FEI VITROBOT MARK III)
3D Reconstruction
Reconstruction MethodSINGLE PARTICLE
Number of Particles36158
Reported Resolution (Å)3.8
Resolution MethodFSC 0.143 CUT-OFF
Other DetailsThe 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 ar ...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 TypeHALF-MAPS REFINED INDEPENDENTLY
Symmetry TypePOINT
Point SymmetryC4
Map-Model Fitting and Refinement
Id1
Refinement SpaceREAL
Refinement ProtocolAB INITIO MODEL
Refinement Target
Overall B Value
Fitting Procedure
DetailsREFINEMENT PROTOCOL--de novo model building DETAILS--The entire ankyrin structure was fitted as a single rigid body. As a result, the first two ankyr ...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.
Data Acquisition
Detector TypeGATAN K2 SUMMIT (4k x 4k)
Electron Dose (electrons/Å**2)21
Imaging Experiment1
Date of Experiment2013-01-01
Temperature (Kelvin)
Microscope ModelFEI POLARA 300
Minimum Defocus (nm)1500
Maximum Defocus (nm)3000
Minimum Tilt Angle (degrees)
Maximum Tilt Angle (degrees)
Nominal CS2
Imaging ModeBRIGHT FIELD
Specimen Holder ModelOTHER
Nominal Magnification31000
Calibrated Magnification31000
SourceFIELD EMISSION GUN
Acceleration Voltage (kV)300
Imaging DetailsGatan K2 Summit operated in super-resolution counting mode, image recorded with dose fractionation method.
EM Software
TaskSoftware PackageVersion
RECONSTRUCTIONRELION
Image Processing
CTF Correction TypeCTF Correction DetailsNumber of Particles SelectedParticle Selection Details
Each particle