EMD-20133

Single-particle
3.0 Å
EMD-20133
3D View Gallery
Deposition: 19/04/2019
Map released: 01/05/2019
Last modified: 20/03/2024
Overview 3D View Sample Experiment Validation Volume Browser Additional data Links
Overview 3D View Sample Experiment Validation Volume Browser Additional data Links

EMD-20133
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3D volume (map.gz) Experimental metadata (xml) Experimental Metadata (cif.gz)
Map-only Validation report (pdf.gz) Map-only Validation report (cif.gz) Map-only Validation report (xml.gz)
Map and PDB model 6on2 summary report (pdf.gz) Map and 6on2 FULL report (pdf.gz)
EMICSS annotations (xml)

Lon Protease from Yersinia pestis with Y2853 substrate

EMD-20133

Single-particle
3.0 Å
EMD-20133
3D View Gallery
Deposition: 19/04/2019
Map released: 01/05/2019
Last modified: 20/03/2024
Overview 3D View Sample Experiment Validation Volume Browser Additional data Links
Method: Single-particle
Aggregation State: Particle
Specimen preparation [1]
Concentration: 0.95 mg/mL
Details: This sample was monodisperse
Buffer
pH: 8.0
Buffer components [5]:
Name Formula Concentration ChEBI
Tris Base Tris 50.0 mM
Potassium Chloride KCl 75.0 mM
Magnesium Chloride MgCl2 10.0 mM
TCEP TCEP 1.0 mM
Adenosine Triphosphate ATP 1.0 mM
Details: Solutions were made fresh from concentrated and filtered using a 0.1 um syringe filter to avoid microbial contamination. Buffers were stored on ice and used within 15 minutes of mixing in order to avoid excess ATP hydrolysis.
Grid
Mesh: 300
Model: Quantifoil, UltrAuFoil, R1.2/1.3
Material: GOLD
Details: Grids were plasma treated for 30 seconds using a 15 mA current operating under atmospheric gases using a glow discharger (Electron Microscopy Sciences).
Pretreatment
Type: GLOW DISCHARGE
Time: {'units': 's', 'valueOf_': '30'}
Atmosphere: AIR
Pressure: 0.01 kPa
Vitrification
Cryogen name: ETHANE
Chamber humidity: 95%
Chamber temperature: 277 K
Instrument: HOMEMADE PLUNGER
Details: 4 uL of sample was applied per grid and manually blotted for 4 seconds followed by immediately plunge-freezing in liquid ethane cooled by liquid nitrogen..
Microscopy [1]
Microscope: FEI TALOS ARCTICA
Illumination mode: FLOOD BEAM
Imaging mode: BRIGHT FIELD
Electron source: FIELD EMISSION GUN
Acceleration voltage: 200 kV
C2 aperture diameter: 70.0 µm
Nominal CS: 2.7 mm
Nominal defocus: 0.8 µm - 1.2 µm
Calibrated defocus: 0.5 µm - 1.5 µm
Nominal magnification: 36000.0
Calibrated magnification: 43478.0
Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Cooling holder cryogen: NITROGEN
Alignment procedure: COMA FREE ( Residual tilt: 0.14 mrad)
Details: Coma-free alignment procedure from Herzik & Wu, Nature Methods (2017). Preliminary grid screening was performed manually prior to data collection.
Temperature
Minimum: 80.0 K
Maximum: 90.0 K
Image Recording [1]
Detector model: GATAN K2 SUMMIT (4k x 4k)
Detector mode: COUNTING
Dimensions: 3710 pixel x 3838 pixel
Frames per image: 0-43
Number of grids: 2
Number of real images: 4071
Average exposure time: 11.0 s
Average electron dose per image: 52.0 e/Å2
Details: Images were collected in counting mode at 4 frames per second
Image processing [1]
Final reconstruction
Resolution: 3.0 Å ( BY AUTHOR)
Resolution method: FSC 0.143 CUT-OFF
Number of classed used: 1
Number of images used: 118143
Algorithm: BACK PROJECTION
⌯ Applied Symmetry
Point group: C1
Software [1]
Name Version Details
RELION 2.0b RELION 2.0b was used to perform final reconstruction
Startup model [1]
Type: INSILICO MODEL
Insilico model: In silico model was created using ab initio 3D reconstruction using a small subset of the data in cryoSPARC
⦨ Initial angle assignment
Type: MAXIMUM LIKELIHOOD
Software [1]
Name Version Details
RELION 2.0b RELION 2.0b was used to assign initial euler angles
⦩ Final angle assignment
Type: MAXIMUM LIKELIHOOD
Details: RELION 2.0b was used to assign initial angles
Software [1]
Name Version Details
RELION 2.0b RELION 2.0b was used to assign final euler angles
Particle selection [1]
Selected Ref. model Method Software Details
1176206 - - - Template-based cross correlation with FindEM
Final 3D classification
Number of classes: 4
Avg. number of members per classes: 75000.0
Details: The final 3D classification had a somewhat asymmetric distribution owing to preferred specimen orientation due to interactions with the air-water interface
Software [1]
Name Version Details
RELION 2.0b RELION 2.0b was used to perform final classification
Map
Format: CCP4
Data type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation details: Composite stitched map for final model building and refinement
⬡ Geometry
X Y Z
Dimensions 130 130 130
Origin 0 0 0
Spacing 130 130 130
Voxel size 1.15 Å 1.15 Å 1.15 Å
Contour list
Primary Level Source
True 0.0403 AUTHOR