+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-12915 | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Title | Cryo-EM structure of T20S proteasome in nanofluidic channels | |||||||||
Map data | Sharpened cryo-EM density | |||||||||
Sample |
| |||||||||
Function / homology | Function and homology information proteasome endopeptidase complex / proteasome core complex, beta-subunit complex / proteasomal protein catabolic process / proteasome core complex, alpha-subunit complex / threonine-type endopeptidase activity / ubiquitin-dependent protein catabolic process / endopeptidase activity / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | Thermoplasma acidophilum (acidophilic) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 5.4 Å | |||||||||
Authors | Huber ST / Sarajlic E / Huijink R / Evers WH / Jakobi AJ | |||||||||
Funding support | European Union, Netherlands, 2 items
| |||||||||
Citation | Journal: Elife / Year: 2022 Title: Nanofluidic chips for cryo-EM structure determination from picoliter sample volumes. Authors: Stefan T Huber / Edin Sarajlic / Roeland Huijink / Felix Weis / Wiel H Evers / Arjen J Jakobi / Abstract: Cryogenic electron microscopy has become an essential tool for structure determination of biological macromolecules. In practice, the difficulty to reliably prepare samples with uniform ice thickness ...Cryogenic electron microscopy has become an essential tool for structure determination of biological macromolecules. In practice, the difficulty to reliably prepare samples with uniform ice thickness still represents a barrier for routine high-resolution imaging and limits the current throughput of the technique. We show that a nanofluidic sample support with well-defined geometry can be used to prepare cryo-EM specimens with reproducible ice thickness from picoliter sample volumes. The sample solution is contained in electron-transparent nanochannels that provide uniform thickness gradients without further optimisation and eliminate the potentially destructive air-water interface. We demonstrate the possibility to perform high-resolution structure determination with three standard protein specimens. Nanofabricated sample supports bear potential to automate the cryo-EM workflow, and to explore new frontiers for cryo-EM applications such as time-resolved imaging and high-throughput screening. #1: Journal: Biorxiv / Year: 2021 Title: Nanofluidic chips for cryo-EM structure determination from picoliter sample volumes Authors: Huber ST / Sarajlic E / Huijink R / Weis F / Evers WH / Jakobi AJ | |||||||||
History |
|
-Structure visualization
Movie |
Movie viewer |
---|---|
Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_12915.map.gz | 59.8 MB | EMDB map data format | |
---|---|---|---|---|
Header (meta data) | emd-12915-v30.xml emd-12915.xml | 20.8 KB 20.8 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_12915_fsc.xml | 8.9 KB | Display | FSC data file |
Images | emd_12915.png | 95.1 KB | ||
Masks | emd_12915_msk_1.map | 64 MB | Mask map | |
Others | emd_12915_additional_1.map.gz emd_12915_additional_2.map.gz emd_12915_half_map_1.map.gz emd_12915_half_map_2.map.gz | 31.7 MB 3.9 MB 59.3 MB 59.3 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-12915 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-12915 | HTTPS FTP |
-Validation report
Summary document | emd_12915_validation.pdf.gz | 708.6 KB | Display | EMDB validaton report |
---|---|---|---|---|
Full document | emd_12915_full_validation.pdf.gz | 708.2 KB | Display | |
Data in XML | emd_12915_validation.xml.gz | 16.2 KB | Display | |
Data in CIF | emd_12915_validation.cif.gz | 21 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-12915 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-12915 | HTTPS FTP |
-Related structure data
Related structure data | 7ohfC C: citing same article (ref.) |
---|---|
Similar structure data | |
EM raw data | EMPIAR-10708 (Title: Apoferritin, TMV and T20S proteasome in nanofluidic channels Data size: 308.3 Data #1: Unaligned multi-frame movies of pyrococcus furiosus apoferritin in silicon nitride nanochannels [micrographs - multiframe] Data #2: Multi-frame movies of T20S proteasome in silicon nitride nanochannels [micrographs - multiframe] Data #3: Multi-frame movies of TMV in silicon nitride nanochannels [micrographs - multiframe]) |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
---|---|
Related items in Molecule of the Month |
-Map
File | Download / File: emd_12915.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Sharpened cryo-EM density | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.288 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
|
-Supplemental data
-Mask #1
File | emd_12915_msk_1.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Projections & Slices |
| ||||||||||||
Density Histograms |
-Additional map: Unsharpened cryo-EM density
File | emd_12915_additional_1.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Unsharpened cryo-EM density | ||||||||||||
Projections & Slices |
| ||||||||||||
Density Histograms |
-Additional map: Local resolution map
File | emd_12915_additional_2.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Local resolution map | ||||||||||||
Projections & Slices |
| ||||||||||||
Density Histograms |
-Half map: Half map A
File | emd_12915_half_map_1.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Half map A | ||||||||||||
Projections & Slices |
| ||||||||||||
Density Histograms |
-Half map: Half map B
File | emd_12915_half_map_2.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Half map B | ||||||||||||
Projections & Slices |
| ||||||||||||
Density Histograms |
-Sample components
-Entire : Thermoplasma acidophilium 20S proteasome
Entire | Name: Thermoplasma acidophilium 20S proteasome |
---|---|
Components |
|
-Supramolecule #1: Thermoplasma acidophilium 20S proteasome
Supramolecule | Name: Thermoplasma acidophilium 20S proteasome / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 |
---|---|
Source (natural) | Organism: Thermoplasma acidophilum (acidophilic) |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Molecular weight | Theoretical: 700 KDa |
-Experimental details
-Structure determination
Method | cryo EM |
---|---|
Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 1.4 mg/mL | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Buffer | pH: 8 Component:
| ||||||||||||
Grid | Model: Homemade / Material: SILICON NITRIDE | ||||||||||||
Vitrification | Cryogen name: ETHANE / Instrument: LEICA PLUNGER Details: The sample was filled into cryoChips through the cantilever and then transferred within ~10 seconds to the Leica plunger for freezing.. | ||||||||||||
Details | The sample was filled into nanofluidic channels. |
-Electron microscopy
Microscope | JEOL 3200FSC |
---|---|
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 121 / Average exposure time: 11.0 sec. / Average electron dose: 59.7 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 4.1 mm |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |