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- EMDB-19791: Flexible reconstruction of a pre-catalytic spliceosome (EMPIAR-10... -

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Basic information

Entry
Database: EMDB / ID: EMD-19791
TitleFlexible reconstruction of a pre-catalytic spliceosome (EMPIAR-10180) using DynaMight
Map datalocal resolution filtered map from the DynaMight half maps
Sample
  • Complex: Pre-catalytic B complex Spliceosome
KeywordsSpliceosome / SPLICING
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 5.98 Å
AuthorsSchwab J / Scheres SHW
Funding support United Kingdom, 1 items
OrganizationGrant numberCountry
UK Research and Innovation (UKRI) United Kingdom
Citation
Journal: Nature / Year: 2017
Title: Structure of a pre-catalytic spliceosome.
Authors: Clemens Plaschka / Pei-Chun Lin / Kiyoshi Nagai /
Abstract: Intron removal requires assembly of the spliceosome on precursor mRNA (pre-mRNA) and extensive remodelling to form the spliceosome's catalytic centre. Here we report the cryo-electron microscopy ...Intron removal requires assembly of the spliceosome on precursor mRNA (pre-mRNA) and extensive remodelling to form the spliceosome's catalytic centre. Here we report the cryo-electron microscopy structure of the yeast Saccharomyces cerevisiae pre-catalytic B complex spliceosome at near-atomic resolution. The mobile U2 small nuclear ribonucleoprotein particle (snRNP) associates with U4/U6.U5 tri-snRNP through the U2/U6 helix II and an interface between U4/U6 di-snRNP and the U2 snRNP SF3b-containing domain, which also transiently contacts the helicase Brr2. The 3' region of the U2 snRNP is flexibly attached to the SF3b-containing domain and protrudes over the concave surface of tri-snRNP, where the U1 snRNP may reside before its release from the pre-mRNA 5' splice site. The U6 ACAGAGA sequence forms a hairpin that weakly tethers the 5' splice site. The B complex proteins Prp38, Snu23 and Spp381 bind the Prp8 N-terminal domain and stabilize U6 ACAGAGA stem-pre-mRNA and Brr2-U4 small nuclear RNA interactions. These results provide important insights into the events leading to active site formation.
#1: Journal: Biorxiv
Title: DynaMight: estimating molecular motions with improved reconstruction from cryo-EM images
Authors: Schwab J / Kimanius D / Burt A / Dendooven T / Scheres SHW
History
DepositionMar 5, 2024-
Header (metadata) releaseMar 13, 2024-
Map releaseMar 13, 2024-
UpdateOct 23, 2024-
Current statusOct 23, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_19791.png

Downloads & links

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Map

FileDownload / File: emd_19791.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationlocal resolution filtered map from the DynaMight half maps
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.7 Å/pix.
x 320 pix.
= 544. Å		1.7 Å/pix.
x 320 pix.
= 544. Å		1.7 Å/pix.
x 320 pix.
= 544. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.7 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.00112
Minimum - Maximum-0.005049644 - 0.009037494
Average (Standard dev.)0.000022444128 (±0.00014893834)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 544.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: unfiltered, unmasked half map reconstructed with DynaMight

Fileemd_19791_half_map_1.map
Annotationunfiltered, unmasked half map reconstructed with DynaMight
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: unfiltered, unmasked half map reconstructed with DynaMight

Fileemd_19791_half_map_2.map
Annotationunfiltered, unmasked half map reconstructed with DynaMight
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Pre-catalytic B complex Spliceosome

EntireName: Pre-catalytic B complex Spliceosome
Components
  • Complex: Pre-catalytic B complex Spliceosome

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Supramolecule #1: Pre-catalytic B complex Spliceosome

SupramoleculeName: Pre-catalytic B complex Spliceosome / type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)

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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: 7.9
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 56.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 5.3 µm / Nominal defocus min: 0.35000000000000003 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 5.98 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 44537
Initial angle assignmentType: PROJECTION MATCHING
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 5.0)

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