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- EMDB-40395: Tertiary structure of an individual particle of self-folding RNA ... -

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Basic information

Entry
Database: EMDB / ID: EMD-40395
TitleTertiary structure of an individual particle of self-folding RNA polymer (particle #161)
Map dataTertiary structure of an individual particle of self-folding RNA polymer (particle #161)
Sample
  • Complex: RNA Origami 6 Helix Bundle with Clasp (6HBC)
    • RNA: RNA Origami 6 Helix Bundle with Clasp (6HBC)
KeywordsSingle molecule structure / Individual Particle cryo-Electron Tomography / IPET / Cryo-ET / tertiary structure / Structural Flexibility / Self-folding mechanism / RNA origami / RNA
Biological speciessynthetic construct (others)
Methodelectron tomography / cryo EM / Resolution: 24.1 Å
AuthorsLiu J / Ren G
Funding support United States, 5 items
OrganizationGrant numberCountry
Department of Energy (DOE, United States)DE-AC02-05CH11231 United States
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)R01HL115153 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM104427 United States
National Institutes of Health/National Institute of Mental Health (NIH/NIMH)R01MH077303 United States
National Institutes of Health/National Institute of Diabetes and Digestive and Kidney Disease (NIH/NIDDK)R01DK042667 United States
CitationJournal: To Be Published
Title: Tertiary Structure of an Individual Particle of Self-folding RNA Polymer
Authors: Liu J / McRae EKS / Zhang M / Geary C / Andersen ES / Ren G
History
DepositionApr 8, 2023-
Header (metadata) releaseApr 10, 2024-
Map releaseApr 10, 2024-
UpdateApr 10, 2024-
Current statusApr 10, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_40395.png

Downloads & links

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Map

FileDownload / File: emd_40395.map.gz / Format: CCP4 / Size: 11.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationTertiary structure of an individual particle of self-folding RNA polymer (particle #161)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.67 Å/pix.
x 144 pix.
= 240.48 Å		1.67 Å/pix.
x 144 pix.
= 240.48 Å		1.67 Å/pix.
x 144 pix.
= 240.48 Å

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.67 Å
Density

Histogram

Histogram (log scale)
Minimum - Maximum-1.0838974 - 3.3949378
Average (Standard dev.)0.042428154 (±0.26454076)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-72-72-72
Dimensions144144144
Spacing144144144
CellA=B=C: 240.48 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : RNA Origami 6 Helix Bundle with Clasp (6HBC)

EntireName: RNA Origami 6 Helix Bundle with Clasp (6HBC)
Components
  • Complex: RNA Origami 6 Helix Bundle with Clasp (6HBC)
    • RNA: RNA Origami 6 Helix Bundle with Clasp (6HBC)

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Supramolecule #1: RNA Origami 6 Helix Bundle with Clasp (6HBC)

SupramoleculeName: RNA Origami 6 Helix Bundle with Clasp (6HBC) / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: The RNA was transcribed from linearized DNA template using T7 RNA polymerase purified in house.
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 230 KDa

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Macromolecule #1: RNA Origami 6 Helix Bundle with Clasp (6HBC)

MacromoleculeName: RNA Origami 6 Helix Bundle with Clasp (6HBC) / type: rna / ID: 1
Source (natural)Organism: synthetic construct (others)
SequenceString: GGGAGAGUAC UAUUCAGAUG CAGACCGCAA GUUCAGAGCG GUUUGCAUCU AGGGUACGUU UUCGAACGUA UCCUCCGACU AAGUGUAUUC GUAUACUUAG UGCCUUGUGC CUGCUUCGGC AGGCAUGACC CAAAUGUGCC UUUCGGGGCA CAUUUCCGGU CAUCCAAGUU ...String:
GGGAGAGUAC UAUUCAGAUG CAGACCGCAA GUUCAGAGCG GUUUGCAUCU AGGGUACGUU UUCGAACGUA UCCUCCGACU AAGUGUAUUC GUAUACUUAG UGCCUUGUGC CUGCUUCGGC AGGCAUGACC CAAAUGUGCC UUUCGGGGCA CAUUUCCGGU CAUCCAAGUU CGCUUGGGUG AUGCGGGCGU AUAGGUUCGU CUAUACGUCC GCGUUUUCCG AGAAGAGGUA ACUCGGGAAA CCGGUCCACG UGACAAAGGU AGAGUUACGU GGAGGGAGCA GCUGCAAAGG GAUAAUGCAG UUGCUGGCUG GAUGCCAGAA CUCACGACUG GCAUCUACGG GGAUGGUGCU CUCCCAAUUC UCCAUUUACC GCCGAAUCGA CCCCAACGUG AGAGGGGUCG GUUCCCCGAG CAUAGACCAA UAUCCCAGGU UUAUGCUCCC CAACGCUGGA CGAACUACCU ACGUCUAGCG UUCCGGCAAA UGAGUCAAUA CCUCAGACUU AUUUGCGGUG CCUGAGCCUA AACUGAACAU GGGUUCAGGC AUCUUGGCUC CAGUUCGCUG GAGCCGACGG UAGCGCUGCG UUCGCGCAGU GCUAGGGAGC AUCCGUUUUC GAGCGGAUGC UGGGCGGUUG CCUGUUCGCA GGCAAUCGGG CCUACUCAUG AUUCGUCAUG AGUGGUGACA GCGUGAUGUU CGCAUUACGC UGUCGGGUAG AUGGAGAAUU

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Experimental details

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Structure determination

Methodcryo EM
Processingelectron tomography
Aggregation stateparticle

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Sample preparation

Concentration0.3 mg/mL
BufferpH: 8
Component:
ConcentrationNameFormula
25.0 mMHEPES
5.0 mMMgCl2MgCl2
100.0 mMKClKCl
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 277 K / Instrument: LEICA EM GP
DetailsIn vitro transcribed RNA was purified by size exclusion chromatography and spin concentrated in amicon spin columns.
SectioningOther: NO SECTIONING

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 2.0 µm / Nominal magnification: 53000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 21 / Average exposure time: 7.39 sec. / Average electron dose: 21.5 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionAlgorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 24.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: IPET
Details: The targeted images of individual particle were reconstructed by Individual-Particle Electron Tomography (IPET). To reduce the missing-wedge artifact caused by the limited tilt angle range, ...Details: The targeted images of individual particle were reconstructed by Individual-Particle Electron Tomography (IPET). To reduce the missing-wedge artifact caused by the limited tilt angle range, the final 3D map was submitted to a low-tilt tomographic 3D reconstruction method (LoTToR). IPET 3D map was low-pass filtered to 0.8 nm following by Gaussian filtering (standard deviation is 3.0) and median filter (3x3x3) using EMAN and UCSF Chimera, and displayed by Chimera with applied the hidden dust function.
Number images used: 21
DetailsMotion correction of the multi-frame movie was conducted by MotionCor2. The tilt series of whole micrographs were initial aligned by IMOD. Additionally, to reduce the image noise, tilt series were further conducted by a machine learning, a median-filter process and a contrast enhancement method.
FSC plot (resolution estimation)

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