Journal: Nat Struct Mol Biol / Year: 2015 Title: Structure of a yeast 40S-eIF1-eIF1A-eIF3-eIF3j initiation complex. Authors: Christopher H S Aylett / Daniel Boehringer / Jan P Erzberger / Tanja Schaefer / Nenad Ban / Abstract: Eukaryotic translation initiation requires cooperative assembly of a large protein complex at the 40S ribosomal subunit. We have resolved a budding yeast initiation complex by cryo-EM, allowing ...Eukaryotic translation initiation requires cooperative assembly of a large protein complex at the 40S ribosomal subunit. We have resolved a budding yeast initiation complex by cryo-EM, allowing placement of prior structures of eIF1, eIF1A, eIF3a, eIF3b and eIF3c. Our structure highlights differences in initiation-complex binding to the ribosome compared to that of mammalian eIF3, demonstrates a direct contact between eIF3j and eIF1A and reveals the network of interactions between eIF3 subunits.
History
Deposition
Dec 18, 2014
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Header (metadata) release
Jan 21, 2015
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Map release
Feb 11, 2015
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Update
Feb 17, 2016
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Current status
Feb 17, 2016
Processing site: PDBe / Status: Released
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Structure visualization
Movie
Surface view with section colored by density value
pH: 7.6 / Details: 25 mM K-HEPES, 10 mM MgCl2, 75 mM KCl, 0.5 mM TCEP
Grid
Details: 200 mesh Quantifoil R 2/2 holey carbon grids with a thin continuous carbon support film
Vitrification
Cryogen name: ETHANE / Chamber humidity: 75 % / Chamber temperature: 120 K / Instrument: HOMEMADE PLUNGER Details: carbon floated on the sample prior to recovery onto the grid and flash-freezing. Method: 5 seconds blotting before plunging.
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Electron microscopy #1
Microscopy ID
1
Microscope
FEI TITAN KRIOS
Temperature
Average: 100 K
Alignment procedure
Legacy - Astigmatism: Objective lens astigmatism was corrected at 150,000 times magnification
Date
Apr 6, 2014
Image recording
Category: CCD / Film or detector model: FEI FALCON II (4k x 4k) / Number real images: 7200 / Average electron dose: 25 e/Å2 Details: Single movie frame readout. 7 frames per exposure. 4 images per hole. Bits/pixel: 16
Electron beam
Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Legacy - Astigmatism: Objective lens astigmatism was corrected at 150,000 times magnification
Date
Jul 16, 2014
Image recording
Category: CCD / Film or detector model: FEI FALCON II (4k x 4k) / Number real images: 8828 / Average electron dose: 25 e/Å2 Details: Single movie frame readout. 7 frames per exposure. Drift corrected in post-processing. 4 images per hole. Bits/pixel: 16
Electron beam
Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
For full details see the supplemental materials and methods in the accompanying publication which provides processing details and the classification schema.
CTF correction
Details: By image
Final reconstruction
Applied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 6.47 Å / Resolution method: OTHER / Software - Name: CTFFIND3, RELION, 1.2 / Number images used: 27354
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