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- EMDB-21531: Head masked 3D refined density of the Cricket Paralysis Virus 5-U... -

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Basic information

Entry
Database: EMDB / ID: EMD-21531
TitleHead masked 3D refined density of the Cricket Paralysis Virus 5-UTR IRES (CrPV 5-UTR-IRES) bound to the small ribosomal subunit used to build the 5-UTR IRES model
Map data
SampleCricket Paralysis Virus 5-UTR IRES (CrPV 5-UTR-IRES) bound to the small ribosomal subunit from a head masked 3D refinement
Biological speciesOryctolagus cuniculus (rabbit)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.79 Å
AuthorsNeupane R / Pisareva V / Rodriguez CF / Pisarev A / Fernandez IS
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM097014 United States
CitationJournal: Elife / Year: 2020
Title: A complex IRES at the 5'-UTR of a viral mRNA assembles a functional 48S complex via an uAUG intermediate.
Authors: Ritam Neupane / Vera P Pisareva / Carlos F Rodriguez / Andrey V Pisarev / Israel S Fernández /
Abstract: Taking control of the cellular apparatus for protein production is a requirement for virus progression. To ensure this control, diverse strategies of cellular mimicry and/or ribosome hijacking have ...Taking control of the cellular apparatus for protein production is a requirement for virus progression. To ensure this control, diverse strategies of cellular mimicry and/or ribosome hijacking have evolved. The initiation stage of translation is especially targeted as it involves multiple steps and the engagement of numerous initiation factors. The use of structured RNA sequences, called nternal ibosomal ntry ites (IRES), in viral RNAs is a widespread strategy for the exploitation of eukaryotic initiation. Using a combination of electron cryo-microscopy (cryo-EM) and reconstituted translation initiation assays with native components, we characterized how a novel IRES at the 5'-UTR of a viral RNA assembles a functional initiation complex via an uAUG intermediate. The IRES features a novel extended, multi-domain architecture, that circles the 40S head. The structures and accompanying functional data illustrate the importance of 5'-UTR regions in translation regulation and underline the relevance of the untapped diversity of viral IRESs.
Validation ReportSummary, Full report, XML, About validation report
History
DepositionMar 8, 2020-
Header (metadata) releaseApr 8, 2020-
Map releaseApr 29, 2020-
UpdateApr 29, 2020-
Current statusApr 29, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.018
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by height
  • Surface level: 0.018
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_21531.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.06 Å/pix.
x 400 pix.
= 424.2 Å
1.06 Å/pix.
x 400 pix.
= 424.2 Å
1.06 Å/pix.
x 400 pix.
= 424.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.0605 Å
Density
Contour LevelBy AUTHOR: 0.018 / Movie #1: 0.018
Minimum - Maximum-0.11528671 - 0.22688793
Average (Standard dev.)0.0000929228 (±0.0032904665)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 424.2 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.06051.06051.0605
M x/y/z400400400
origin x/y/z0.0000.0000.000
length x/y/z424.200424.200424.200
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ350350350
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS400400400
D min/max/mean-0.1150.2270.000

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Supplemental data

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Segmentation: #1

Fileemd_21531_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: Unsharpened map before post-processing for head masked

Fileemd_21531_additional.map
AnnotationUnsharpened map before post-processing for head masked
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: Half map 1 for head masked

Fileemd_21531_half_map_1.map
AnnotationHalf map 1 for head masked
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: Half map 2 for head masked

Fileemd_21531_half_map_2.map
AnnotationHalf map 2 for head masked
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire Cricket Paralysis Virus 5-UTR IRES (CrPV 5-UTR-IRES) bound to the...

EntireName: Cricket Paralysis Virus 5-UTR IRES (CrPV 5-UTR-IRES) bound to the small ribosomal subunit from a head masked 3D refinement
Number of components: 1

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Component #1: protein, Cricket Paralysis Virus 5-UTR IRES (CrPV 5-UTR-IRES) bou...

ProteinName: Cricket Paralysis Virus 5-UTR IRES (CrPV 5-UTR-IRES) bound to the small ribosomal subunit from a head masked 3D refinement
Recombinant expression: No
SourceSpecies: Oryctolagus cuniculus (rabbit)

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionpH: 7.5
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 277.15 K / Humidity: 100 %
Details: Grids were blotted for 2.5s and flash cooled in liquid ethane.

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 56.9 e/Å2 / Illumination mode: FLOOD BEAM
LensImaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C1 (asymmetric) / Number of projections: 37701
Details: The microscope was equipped with an energy filter with slits aperture of 20eV, installed before the detector.
3D reconstructionSoftware: RELION / Resolution: 2.79 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution estimation)

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