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- EMDB-3769: Cryo-EM structure of a pre-catalytic human spliceosome primed for... -

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Basic information

Entry
Database: EMDB / ID: EMD-3769
TitleCryo-EM structure of a pre-catalytic human spliceosome primed for activation (B complex). Unmasked refinement, NON-sharpened map
Map dataNON-sharpened, unmasked refinement. Map was aligned to high resolution map (EMDB 3766) and RESAMPLED from 256 px boxsize to 432 px boxsize in Chimera to fit high-resolution map.
Sample
  • Complex: Cryo-EM structure of a pre-catalytic human spliceosome primed for activation (B complex). Unmasked refinement, NON-sharpened map
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 9.9 Å
AuthorsBertram K
CitationJournal: Cell / Year: 2017
Title: Cryo-EM Structure of a Pre-catalytic Human Spliceosome Primed for Activation.
Authors: Karl Bertram / Dmitry E Agafonov / Olexandr Dybkov / David Haselbach / Majety N Leelaram / Cindy L Will / Henning Urlaub / Berthold Kastner / Reinhard Lührmann / Holger Stark /
Abstract: Little is known about the spliceosome's structure before its extensive remodeling into a catalytically active complex. Here, we report a 3D cryo-EM structure of a pre-catalytic human spliceosomal B ...Little is known about the spliceosome's structure before its extensive remodeling into a catalytically active complex. Here, we report a 3D cryo-EM structure of a pre-catalytic human spliceosomal B complex. The U2 snRNP-containing head domain is connected to the B complex main body via three main bridges. U4/U6.U5 tri-snRNP proteins, which are located in the main body, undergo significant rearrangements during tri-snRNP integration into the B complex. These include formation of a partially closed Prp8 conformation that creates, together with Dim1, a 5' splice site (ss) binding pocket, displacement of Sad1, and rearrangement of Brr2 such that it contacts its U4/U6 substrate and is poised for the subsequent spliceosome activation step. The molecular organization of several B-specific proteins suggests that they are involved in negatively regulating Brr2, positioning the U6/5'ss helix, and stabilizing the B complex structure. Our results indicate significant differences between the early activation phase of human and yeast spliceosomes.
History
DepositionJun 20, 2017-
Header (metadata) releaseAug 16, 2017-
Map releaseAug 16, 2017-
UpdateAug 23, 2017-
Current statusAug 23, 2017Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.016378257
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.016378257
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_3769.png

Downloads & links

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Map

FileDownload / File: emd_3769.map.gz / Format: CCP4 / Size: 307.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationNON-sharpened, unmasked refinement. Map was aligned to high resolution map (EMDB 3766) and RESAMPLED from 256 px boxsize to 432 px boxsize in Chimera to fit high-resolution map.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.16 Å/pix.
x 432 pix.
= 501.12 Å		1.16 Å/pix.
x 432 pix.
= 501.12 Å		1.16 Å/pix.
x 432 pix.
= 501.12 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.16 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.04 / Movie #1: 0.0163783
Minimum - Maximum-0.022836115 - 0.07936885
Average (Standard dev.)-0.00015294988 (±0.00620588)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions432432432
Spacing432432432
CellA=B=C: 501.12 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.161.161.16
M x/y/z432432432
origin x/y/z0.0000.0000.000
length x/y/z501.120501.120501.120
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS432432432
D min/max/mean-0.0230.079-0.000

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Supplemental data

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Additional map: NON-sharpened, unmasked refinement. Map with original boxsize used...

Fileemd_3769_additional.map
AnnotationNON-sharpened, unmasked refinement. Map with original boxsize used in refinement
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half-map 1

Fileemd_3769_half_map_1.map
Annotationhalf-map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half-map 2

Fileemd_3769_half_map_2.map
Annotationhalf-map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Cryo-EM structure of a pre-catalytic human spliceosome primed for...

EntireName: Cryo-EM structure of a pre-catalytic human spliceosome primed for activation (B complex). Unmasked refinement, NON-sharpened map
Components
  • Complex: Cryo-EM structure of a pre-catalytic human spliceosome primed for activation (B complex). Unmasked refinement, NON-sharpened map

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Supramolecule #1: Cryo-EM structure of a pre-catalytic human spliceosome primed for...

SupramoleculeName: Cryo-EM structure of a pre-catalytic human spliceosome primed for activation (B complex). Unmasked refinement, NON-sharpened map
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#43
Source (natural)Organism: Homo sapiens (human)

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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 / Chamber humidity: 80 % / Chamber temperature: 277 K

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Digitization - Sampling interval: 14.0 µm / Digitization - Frames/image: 2-17 / Average exposure time: 0.05 sec. / Average electron dose: 1.5 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD / Cs: 0.01 mm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

DetailsFALCON III Direct Electron Detector
Startup modelType of model: OTHER / Details: Reconstruction from negatively stained particles
Final reconstructionResolution.type: BY AUTHOR / Resolution: 9.9 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 44629
Initial angle assignmentType: COMMON LINE
Final angle assignmentType: OTHER

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Atomic model buiding 1

RefinementSpace: REAL

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