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- EMDB-2276: Ribosome structures to near-atomic resolution from thirty thousan... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-2276 | |||||||||
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Title | Ribosome structures to near-atomic resolution from thirty thousand cryo-EM particles | |||||||||
![]() | Reconstruction of S.cereviseae 80S ribosome (conformation 2) | |||||||||
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![]() | cryo-EM / single-particle analysis / direct electron detectors / ribosome / RELION / statistical movie processing | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.6 Å | |||||||||
![]() | Bai XC / Fernandez IS / McMullan G / Scheres SHW | |||||||||
![]() | ![]() Title: Ribosome structures to near-atomic resolution from thirty thousand cryo-EM particles. Authors: Xiao-Chen Bai / Israel S Fernandez / Greg McMullan / Sjors H W Scheres / ![]() Abstract: Although electron cryo-microscopy (cryo-EM) single-particle analysis has become an important tool for structural biology of large and flexible macro-molecular assemblies, the technique has not yet ...Although electron cryo-microscopy (cryo-EM) single-particle analysis has become an important tool for structural biology of large and flexible macro-molecular assemblies, the technique has not yet reached its full potential. Besides fundamental limits imposed by radiation damage, poor detectors and beam-induced sample movement have been shown to degrade attainable resolutions. A new generation of direct electron detectors may ameliorate both effects. Apart from exhibiting improved signal-to-noise performance, these cameras are also fast enough to follow particle movements during electron irradiation. Here, we assess the potentials of this technology for cryo-EM structure determination. Using a newly developed statistical movie processing approach to compensate for beam-induced movement, we show that ribosome reconstructions with unprecedented resolutions may be calculated from almost two orders of magnitude fewer particles than used previously. Therefore, this methodology may expand the scope of high-resolution cryo-EM to a broad range of biological specimens.DOI:http://dx.doi.org/10.7554/eLife.00461.001. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 49.4 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 11.5 KB 11.5 KB | Display Display | ![]() |
Images | ![]() | 279.2 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
File | ![]() | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Reconstruction of S.cereviseae 80S ribosome (conformation 2) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.77 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : S.cereviseae 80S ribosome
Entire | Name: S.cereviseae 80S ribosome |
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Components |
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-Supramolecule #1000: S.cereviseae 80S ribosome
Supramolecule | Name: S.cereviseae 80S ribosome / type: sample / ID: 1000 / Number unique components: 1 |
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Molecular weight | Experimental: 4.2 MDa / Theoretical: 4.2 MDa |
-Supramolecule #1: 80S ribosome
Supramolecule | Name: 80S ribosome / type: complex / ID: 1 / Recombinant expression: No / Ribosome-details: ribosome-eukaryote: ALL |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Experimental: 4.2 MDa / Theoretical: 4.2 MDa |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.3 mg/mL |
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Buffer | pH: 7.45 Details: 3mM Hepes-KOH, 6.6 mM Tris-acetate pH 7.2, 3 mM NH4Cl, 6.6 mM NH4-acetate, 48 mM K-acetate, 4 mM Mg-acetate, 2.4 mM DTT |
Grid | Details: Quantifoil grids (2/2) with 3 nm thin carbon on top |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 90 K / Instrument: FEI VITROBOT MARK II / Method: Blot 2.5 seconds before plunging |
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Electron microscopy
Microscope | FEI POLARA 300 |
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Temperature | Min: 80 K / Max: 90 K / Average: 85 K |
Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism was corrected at 59,000 times magnification |
Date | Jul 15, 2012 |
Image recording | Category: CCD / Film or detector model: FEI FALCON I (4k x 4k) / Digitization - Sampling interval: 14 µm / Number real images: 200 / Average electron dose: 16 e/Å2 Details: Every image is the average of sixteen frames recorded by the direct electron detector |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Calibrated magnification: 79096 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2 mm / Nominal defocus max: 3.44 µm / Nominal defocus min: 1.191 µm / Nominal magnification: 59000 |
Sample stage | Specimen holder model: GATAN LIQUID NITROGEN |
Experimental equipment | ![]() Model: Tecnai Polara / Image courtesy: FEI Company |
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Image processing
Details | Use a newly developed statistical movie processing to compensate for beam-induced movement |
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CTF correction | Details: each image |
Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 4.6 Å / Resolution method: OTHER / Software - Name: CTFFIND3, RELION Details: Use a newly developed statistical movie processing approach to compensate for beam-induced movement Number images used: 22638 |
-Atomic model buiding 1
Initial model | PDB ID: ![]() 3u5b |
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Software | Name: ![]() |
Details | Protocol: Rigid body. The domains were separately fitted by manual docking using program Chimera |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |
-Atomic model buiding 2
Initial model | PDB ID: ![]() 3u5c |
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Software | Name: ![]() |
Details | Protocol: Rigid body. The domains were separately fitted by manual docking using program Chimera |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |
-Atomic model buiding 3
Initial model | PDB ID: ![]() 3u5d |
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Software | Name: ![]() |
Details | Protocol: Rigid body. The domains were separately fitted by manual docking using program Chimera |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |
-Atomic model buiding 4
Initial model | PDB ID: ![]() 3u5e |
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Software | Name: ![]() |
Details | Protocol: Rigid body. The domains were separately fitted by manual docking using program Chimera |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |