5GAQ

Cryo-EM structure of the Lysenin Pore


ELECTRON MICROSCOPY

Refinement

RMS Deviations
KeyRefinement Restraint Deviation
r_dihedral_angle_2_deg27.002
r_long_range_B_refined25.055
r_long_range_B_other25.055
r_scangle_other19.592
r_mcangle_it13.252
r_mcangle_other13.252
r_scbond_it11.681
r_scbond_other11.68
r_dihedral_angle_3_deg11.634
r_mcbond_it9.124
RMS Deviations
KeyRefinement Restraint Deviation
r_dihedral_angle_2_deg27.002
r_long_range_B_refined25.055
r_long_range_B_other25.055
r_scangle_other19.592
r_mcangle_it13.252
r_mcangle_other13.252
r_scbond_it11.681
r_scbond_other11.68
r_dihedral_angle_3_deg11.634
r_mcbond_it9.124
r_mcbond_other9.124
r_dihedral_angle_4_deg8.954
r_dihedral_angle_1_deg5.564
r_angle_refined_deg1.063
r_angle_other_deg0.792
r_chiral_restr0.067
r_bond_refined_d0.005
r_gen_planes_refined0.003
r_bond_other_d0.002
r_gen_planes_other0.001
r_nbd_refined
r_nbd_other
r_nbtor_refined
r_nbtor_other
r_xyhbond_nbd_refined
r_xyhbond_nbd_other
r_metal_ion_refined
r_metal_ion_other
r_symmetry_vdw_refined
r_symmetry_vdw_other
r_symmetry_hbond_refined
r_symmetry_hbond_other
r_symmetry_metal_ion_refined
r_symmetry_metal_ion_other
r_scangle_it
r_rigid_bond_restr
r_sphericity_free
r_sphericity_bonded
Sample
Membrane-inserted form of the nonameric pore forming protein Lysenin
Specimen Preparation
Sample Aggregation StatePARTICLE
Vitrification Instrument
Cryogen NameETHANE
Sample Vitrification Details
3D Reconstruction
Reconstruction MethodSINGLE PARTICLE
Number of Particles29329
Reported Resolution (Å)3.1
Resolution MethodFSC 0.143 CUT-OFF
Other Details
Refinement Type
Symmetry TypePOINT
Point SymmetryC9
Map-Model Fitting and Refinement
Id1 (3ZXD)
Refinement SpaceREAL
Refinement ProtocolAB INITIO MODEL
Refinement Target
Overall B Value
Fitting Procedure
DetailsInitial docking of C-terminal region of lysenin monomer to EM map was performed using UCSF Chimera followed by model N-terminal extension in COOT.
Data Acquisition
Detector TypeGATAN K2 SUMMIT (4k x 4k)
Electron Dose (electrons/Å**2)47
Imaging Experiment1
Date of Experiment
Temperature (Kelvin)
Microscope ModelFEI TITAN KRIOS
Minimum Defocus (nm)1000
Maximum Defocus (nm)4500
Minimum Tilt Angle (degrees)
Maximum Tilt Angle (degrees)
Nominal CS2.7
Imaging ModeBRIGHT FIELD
Specimen Holder ModelFEI TITAN KRIOS AUTOGRID HOLDER
Nominal Magnification81000
Calibrated Magnification34965
SourceFIELD EMISSION GUN
Acceleration Voltage (kV)300
Imaging DetailsDirect alignments: Beam tilt pivot points, Beam shift, Comma Free. C2 aperture centering, C2 lens astigmatism correction. Objective aperture centering and objective lens astigmatism correction. Energy filter Tuning and occasional ZLP centering.
EM Software
TaskSoftware PackageVersion
PARTICLE SELECTIONRELION1.4
IMAGE ACQUISITIONTitan User Interface
CTF CORRECTIONGctf
MODEL FITTINGCoot0.8.2
INITIAL EULER ASSIGNMENTRELION1.4
FINAL EULER ASSIGNMENTRELION1.4
CLASSIFICATIONRELION1.4
RECONSTRUCTIONRELION1.4
MODEL REFINEMENTREFMAC5
Image Processing
CTF Correction TypeCTF Correction DetailsNumber of Particles SelectedParticle Selection Details
PHASE FLIPPING AND AMPLITUDE CORRECTION53779Autopicking performed with RELION after manually picking 1000 particles to create 2D Classes (references) for autopicking.