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- EMDB-23401: Cryo-EM structure of the B dENE construct complexed with a 28-mer... -

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Entry
Database: EMDB / ID: EMD-23401
TitleCryo-EM structure of the B dENE construct complexed with a 28-mer poly(A)
Map dataCryo-EM structure of the B dENE construct complexed with a 28-mer poly(A).
Sample
  • Complex: dENE construct complexed with a 28-mer poly(A)
    • RNA: B dENE construct
    • RNA: poly(A)
KeywordsRNA triple helix / RNA stability / poly(A) / SAXS and cryo-EM / RNA
Biological speciesOryza sativa (Asian cultivated rice) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 5.6 Å
AuthorsTorabi S / Chen Y
Funding support United States, 3 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)P41GM103832 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)P01AI120943 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)S10OD021600 United States
CitationJournal: Proc Natl Acad Sci U S A / Year: 2021
Title: Structural analyses of an RNA stability element interacting with poly(A).
Authors: Seyed-Fakhreddin Torabi / Yen-Lin Chen / Kaiming Zhang / Jimin Wang / Suzanne J DeGregorio / Anand T Vaidya / Zhaoming Su / Suzette A Pabit / Wah Chiu / Lois Pollack / Joan A Steitz /
Abstract: Cis-acting RNA elements are crucial for the regulation of polyadenylated RNA stability. The element for nuclear expression (ENE) contains a U-rich internal loop flanked by short helices. An ENE ...Cis-acting RNA elements are crucial for the regulation of polyadenylated RNA stability. The element for nuclear expression (ENE) contains a U-rich internal loop flanked by short helices. An ENE stabilizes RNA by sequestering the poly(A) tail via formation of a triplex structure that inhibits a rapid deadenylation-dependent decay pathway. Structure-based bioinformatic studies identified numerous ENE-like elements in evolutionarily diverse genomes, including a subclass containing two ENE motifs separated by a short double-helical region (double ENEs [dENEs]). Here, the structure of a dENE derived from a rice transposable element (TWIFB1) before and after poly(A) binding (∼24 kDa and ∼33 kDa, respectively) is investigated. We combine biochemical structure probing, small angle X-ray scattering (SAXS), and cryo-electron microscopy (cryo-EM) to investigate the dENE structure and its local and global structural changes upon poly(A) binding. Our data reveal 1) the directionality of poly(A) binding to the dENE, and 2) that the dENE-poly(A) interaction involves a motif that protects the 3'-most seven adenylates of the poly(A). Furthermore, we demonstrate that the dENE does not undergo a dramatic global conformational change upon poly(A) binding. These findings are consistent with the recently solved crystal structure of a dENE+poly(A) complex [S.-F. Torabi , 371, eabe6523 (2021)]. Identification of additional modes of poly(A)-RNA interaction opens new venues for better understanding of poly(A) tail biology.
History
DepositionFeb 1, 2021-
Header (metadata) releaseApr 14, 2021-
Map releaseApr 14, 2021-
UpdateMar 6, 2024-
Current statusMar 6, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.459
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.459
  • Imaged by UCSF Chimera
  • Download
  • Surface view with fitted model
  • Atomic models: PDB-7ljy
  • Surface level: 0.459
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_23401.png

Downloads & links

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Map

FileDownload / File: emd_23401.map.gz / Format: CCP4 / Size: 42.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryo-EM structure of the B dENE construct complexed with a 28-mer poly(A).
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.82 Å/pix.
x 224 pix.
= 183.68 Å		0.82 Å/pix.
x 224 pix.
= 183.68 Å		0.82 Å/pix.
x 224 pix.
= 183.68 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.82 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.459 / Movie #1: 0.459
Minimum - Maximum-3.6010506 - 3.1136997
Average (Standard dev.)0.0020915293 (±0.06452999)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions224224224
Spacing224224224
CellA=B=C: 183.68 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.820.820.82
M x/y/z224224224
origin x/y/z0.0000.0000.000
length x/y/z183.680183.680183.680
α/β/γ90.00090.00090.000
start NX/NY/NZ645365
NX/NY/NZ139143124
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS224224224
D min/max/mean-3.6013.1140.002

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Supplemental data

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Sample components

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Entire : dENE construct complexed with a 28-mer poly(A)

EntireName: dENE construct complexed with a 28-mer poly(A)
Components
  • Complex: dENE construct complexed with a 28-mer poly(A)
    • RNA: B dENE construct
    • RNA: poly(A)

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Supramolecule #1: dENE construct complexed with a 28-mer poly(A)

SupramoleculeName: dENE construct complexed with a 28-mer poly(A) / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Oryza sativa (Asian cultivated rice)
Molecular weightTheoretical: 33 KDa

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Macromolecule #1: B dENE construct

MacromoleculeName: B dENE construct / type: rna / ID: 1 / Number of copies: 1
Source (natural)Organism: Oryza sativa (Asian cultivated rice)
Molecular weightTheoretical: 24.242186 KDa
SequenceString:
GGGUACUCUU UUCUUUGUCA UGGUUUUCUC AGGCGAAAGU CUGAGUUUUU ACAUGACAAA GUUUUUAACG AGGCCC

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Macromolecule #2: poly(A)

MacromoleculeName: poly(A) / type: rna / ID: 2 / Number of copies: 1
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 9.172806 KDa
SequenceString:
AAAAAAAAAA AAAAAAAAAA AAAAAAAA

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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
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 40 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 %

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 5.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 2.15.0) / Number images used: 283486
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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Atomic model buiding 1

RefinementProtocol: RIGID BODY FIT
Output model

PDB-7ljy:
Cryo-EM structure of the B dENE construct complexed with a 28-mer poly(A)

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