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- EMDB-35233: Yeast 40S ribosomal subunit -

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Basic information

Entry
Database: EMDB / ID: EMD-35233
TitleYeast 40S ribosomal subunit
Map dataCryo-EM map of yeast 40S ribosomal subunit
Sample
  • Complex: 40S subunit of yeast
KeywordsYeast small ribosomal subunit 40S / RIBOSOME
Biological speciesKluyveromyces lactis NRRL Y-1140 (yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.0 Å
AuthorsDatey A / Khaja FT / Hussain T
Funding support India, 1 items
OrganizationGrant numberCountry
Department of Biotechnology (DBT, India)IA/I/17/2/503313 India
CitationJournal: Biochemistry / Year: 2025
Title: Yeast Eukaryotic Initiation Factor 4B Remodels the MRNA Entry Site on the Small Ribosomal Subunit.
Authors: Ayushi Datey / Prafful Sharma / Faisal Tarique Khaja / Huma Rahil / Tanweer Hussain /
Abstract: Eukaryotic Initiation Factor 4 (eIF4) is a group of factors that activates mRNA for translation and recruit 43S preinitiation complex (PIC) to the mRNA 5' end, forming the 48S PIC. The eIF4 factors ...Eukaryotic Initiation Factor 4 (eIF4) is a group of factors that activates mRNA for translation and recruit 43S preinitiation complex (PIC) to the mRNA 5' end, forming the 48S PIC. The eIF4 factors include mRNA 5' cap-binding protein eIF4E, ATP-dependent RNA helicase eIF4A, and scaffold protein eIF4G, which anchors eIF4A and eIF4E. Another eIF4 factor, eIF4B, stimulates the RNA helicase activity of eIF4A and facilitates mRNA recruitment. However, the mechanisms by which eIF4B binds the 40S ribosomal subunit and promotes mRNA recruitment remain poorly understood. Using cryo-Eletron Microscopy (cryo-EM), we obtained a map of the yeast 40S ribosomal subunit in a complex with eIF4B (40S-eIF4B complex). An extra density, tentatively assigned to yeast eIF4B, was observed near the mRNA entry channel of the 40S, contacting ribosomal proteins uS10, uS3, and eS10 as well as rRNA helix h16. Predictive modeling of the 40S-eIF4B complex suggests that the N-terminal domain of eIF4B binds near the mRNA entry channel, overlapping with the extra density observed in the 40S-eIF4B map. The partially open conformation of 40S in the 40S-eIF4B map is incompatible with eIF3j binding observed in the 48S PIC. Additionally, the extra density at the mRNA entry channel poses steric hindrance for eIF3g binding in the 48S PIC. Thus, structural insights suggest that eIF4B facilitates the release of eIF3j and the relocation of the eIF3b-g-i module during mRNA recruitment, thereby advancing our understanding of eIF4B's role in translation initiation.
History
DepositionFeb 2, 2023-
Header (metadata) releaseFeb 7, 2024-
Map releaseFeb 7, 2024-
UpdateFeb 5, 2025-
Current statusFeb 5, 2025Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_35233.png

Downloads & links

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Map

FileDownload / File: emd_35233.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryo-EM map of yeast 40S ribosomal subunit
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.17 Å/pix.
x 360 pix.
= 421.2 Å		1.17 Å/pix.
x 360 pix.
= 421.2 Å		1.17 Å/pix.
x 360 pix.
= 421.2 Å

Surface

Projections

Slices (1/3)

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.17 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.006
Minimum - Maximum-0.033786345 - 0.08014698
Average (Standard dev.)0.000085198684 (±0.0026404974)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 421.19998 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Auto-refined map of yeast 40S ribosomal subunit.

Fileemd_35233_additional_1.map
AnnotationAuto-refined map of yeast 40S ribosomal subunit.
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map 1 of yeast 40S ribosomal subunit

Fileemd_35233_half_map_1.map
AnnotationHalf map 1 of yeast 40S ribosomal subunit
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map 2 of yeast 40S ribosomal subunit

Fileemd_35233_half_map_2.map
AnnotationHalf map 2 of yeast 40S ribosomal subunit
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : 40S subunit of yeast

EntireName: 40S subunit of yeast
Components
  • Complex: 40S subunit of yeast

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Supramolecule #1: 40S subunit of yeast

SupramoleculeName: 40S subunit of yeast / type: complex / ID: 1 / Parent: 0
Details: This 40S cryo-EM map was obtained during masked and unmasked 3D classification of the cryo-EM dataset of the 40S-eIF4B complex
Source (natural)Organism: Kluyveromyces lactis NRRL Y-1140 (yeast) / Strain: NRRL Y-1140

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.4
Component:
ConcentrationFormulaName
20.0 mMC8H18N2O4SHEPES
100.0 mMCH3CO2KPotassium acetate
2.5 mMC2H6O4MgMagnesium acetate
2.0 mMC4H10O2S2Dithiothreitol

Details: 20 mM HEPES pH7.4, 100 mM potassium acetate pH7.6, 2.5 mM Magnesium acetate, 2 mM DTT
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 90 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 100.0 kPa
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK IV
Details: Vitrification was performed in nitrogen atmosphere.
DetailsThis class of empty or only 40S was a part of the reaction mixture containing 40S and the Saccharomyces cerevisiae eukaryotic initiation factor 4B present in a 1:10 molar ratio

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
TemperatureMin: 77.0 K / Max: 98.0 K
DetailsPreliminary grid screening was done manually using the Latitude-S
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 4-16 / Number grids imaged: 6 / Number real images: 7975 / Average exposure time: 8.0 sec. / Average electron dose: 50.0 e/Å2
Details: Images were collected in movie-mode at 20 frames per 8 seconds
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Calibrated defocus max: 3.0 µm / Calibrated defocus min: 1.5 µm / Calibrated magnification: 42000 / Illumination mode: OTHER / Imaging mode: DIFFRACTION / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 42000
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1492547
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

3jam


Details: map generated from 3JAM was low-pass filtered and used as a reference for 3D autorefinement
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 4.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1.1) / Number images used: 465216
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1.1)
Final 3D classificationNumber classes: 2 / Software - Name: RELION (ver. 3.1.1)
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementProtocol: RIGID BODY FIT / Target criteria: Cross-correlation coefficient

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