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- EMDB-31332: The cryo-EM map of the DDX42-SF3b complex in N-terminus of DDX42 -

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Basic information

Entry
Database: EMDB / ID: EMD-31332
TitleThe cryo-EM map of the DDX42-SF3b complex in N-terminus of DDX42
Map dataThe EM map of DDX42 Nhelix region
Sample
  • Complex: The DDX42-SF3b core complex
KeywordsDDX32 / SF3B1 / SF3b / U2 snRNP / SPLICING
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.7 Å
AuthorsZhang X / Zhan X / Shi Y
Funding support China, 1 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)31930059 China
CitationJournal: Nat Commun / Year: 2023
Title: Mechanisms of the RNA helicases DDX42 and DDX46 in human U2 snRNP assembly.
Authors: Fenghua Yang / Tong Bian / Xiechao Zhan / Zhe Chen / Zhihan Xing / Nicolas A Larsen / Xiaofeng Zhang / Yigong Shi /
Abstract: Three RNA helicases - DDX42, DDX46 and DHX15 - are found to be associated with human U2 snRNP, but their roles and mechanisms in U2 snRNP and spliceosome assembly are insufficiently understood. Here ...Three RNA helicases - DDX42, DDX46 and DHX15 - are found to be associated with human U2 snRNP, but their roles and mechanisms in U2 snRNP and spliceosome assembly are insufficiently understood. Here we report the cryo-electron microscopy (cryo-EM) structures of the DDX42-SF3b complex and a putative assembly precursor of 17S U2 snRNP that contains DDX42 (DDX42-U2 complex). DDX42 is anchored on SF3B1 through N-terminal sequences, with its N-plug occupying the RNA path of SF3B1. The binding mode of DDX42 to SF3B1 is in striking analogy to that of DDX46. In the DDX42-U2 complex, the N-terminus of DDX42 remains anchored on SF3B1, but the helicase domain has been displaced by U2 snRNA and TAT-SF1. Through in vitro assays, we show DDX42 and DDX46 are mutually exclusive in terms of binding to SF3b. Cancer-driving mutations of SF3B1 target the residues in the RNA path that directly interact with DDX42 and DDX46. These findings reveal the distinct roles of DDX42 and DDX46 in assembly of 17S U2 snRNP and provide insights into the mechanisms of SF3B1 cancer mutations.
History
DepositionMay 21, 2021-
Header (metadata) releaseAug 3, 2022-
Map releaseAug 3, 2022-
UpdateAug 16, 2023-
Current statusAug 16, 2023Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_31332.png

Downloads & links

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Map

FileDownload / File: emd_31332.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationThe EM map of DDX42 Nhelix region
Voxel sizeX=Y=Z: 1.087 Å
Density
Contour LevelBy AUTHOR: 0.04
Minimum - Maximum-0.09485854 - 0.17972563
Average (Standard dev.)0.00009876591 (±0.0050358106)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions240240240
Spacing240240240
CellA=B=C: 260.88 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : The DDX42-SF3b core complex

EntireName: The DDX42-SF3b core complex
Components
  • Complex: The DDX42-SF3b core complex

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Supramolecule #1: The DDX42-SF3b core complex

SupramoleculeName: The DDX42-SF3b core complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#5
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

Concentration1.5 mg/mL
BufferpH: 7.9
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: EMDB MAP
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionAlgorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 2.7 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 104710

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