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- EMDB-24284: Azoarcus group I intron, symmetry-expanded monomer from a synthet... -

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Basic information

Entry
Database: EMDB / ID: EMD-24284
TitleAzoarcus group I intron, symmetry-expanded monomer from a synthetic trimeric construct
Map dataAzoarcus group I intron, symmetry-expanded monomer from a synthetic trimeric construct
Sample
  • Complex: Azoarcus group I intron, symmetry-expanded monomer from a synthetic trimeric construct
    • RNA: Azoarcus group I intron
    • Other: Ligated exon mimic of the Azoarcus Group I intron
Biological speciesAzoarcus olearius (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.9 Å
AuthorsThelot F / Liu D / Liao M / Yin P
Funding support United States, 5 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM122797 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)5DP1GM133052 United States
National Science Foundation (NSF, United States)CMMI-1333215 United States
National Science Foundation (NSF, United States)CMMI-1344915 United States
National Science Foundation (NSF, United States)CBET-1729397 United States
CitationJournal: Nat Methods / Year: 2022
Title: Sub-3-Å cryo-EM structure of RNA enabled by engineered homomeric self-assembly.
Authors: Di Liu / François A Thélot / Joseph A Piccirilli / Maofu Liao / Peng Yin /
Abstract: High-resolution structural studies are essential for understanding the folding and function of diverse RNAs. Herein, we present a nanoarchitectural engineering strategy for efficient structural ...High-resolution structural studies are essential for understanding the folding and function of diverse RNAs. Herein, we present a nanoarchitectural engineering strategy for efficient structural determination of RNA-only structures using single-particle cryogenic electron microscopy (cryo-EM). This strategy-ROCK (RNA oligomerization-enabled cryo-EM via installing kissing loops)-involves installing kissing-loop sequences onto the functionally nonessential stems of RNAs for homomeric self-assembly into closed rings with multiplied molecular weights and mitigated structural flexibility. ROCK enables cryo-EM reconstruction of the Tetrahymena group I intron at 2.98-Å resolution overall (2.85 Å for the core), allowing de novo model building of the complete RNA, including the previously unknown peripheral domains. ROCK is further applied to two smaller RNAs-the Azoarcus group I intron and the FMN riboswitch, revealing the conformational change of the former and the bound ligand in the latter. ROCK holds promise to greatly facilitate the use of cryo-EM in RNA structural studies.
History
DepositionJun 22, 2021-
Header (metadata) releaseMay 4, 2022-
Map releaseMay 4, 2022-
UpdateJun 1, 2022-
Current statusJun 1, 2022Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_24284.png

Downloads & links

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Map

FileDownload / File: emd_24284.map.gz / Format: CCP4 / Size: 5.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationAzoarcus group I intron, symmetry-expanded monomer from a synthetic trimeric construct
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.23 Å/pix.
x 61 pix.
= 75.03 Å		1.23 Å/pix.
x 162 pix.
= 199.26 Å		1.23 Å/pix.
x 147 pix.
= 180.81 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX=Y=Z: 1.23 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.182
Minimum - Maximum-1.2116393 - 1.422212
Average (Standard dev.)7.281152e-13 (±0.071785115)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-67-71-31
Dimensions16214761
Spacing14716261
CellA: 180.81 Å / B: 199.26001 Å / C: 75.03 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : Azoarcus group I intron, symmetry-expanded monomer from a synthet...

EntireName: Azoarcus group I intron, symmetry-expanded monomer from a synthetic trimeric construct
Components
  • Complex: Azoarcus group I intron, symmetry-expanded monomer from a synthetic trimeric construct
    • RNA: Azoarcus group I intron
    • Other: Ligated exon mimic of the Azoarcus Group I intron

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Supramolecule #1: Azoarcus group I intron, symmetry-expanded monomer from a synthet...

SupramoleculeName: Azoarcus group I intron, symmetry-expanded monomer from a synthetic trimeric construct
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Azoarcus olearius (bacteria)
Molecular weightTheoretical: 80 KDa

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Macromolecule #1: Azoarcus group I intron

MacromoleculeName: Azoarcus group I intron / type: rna / ID: 1
Source (natural)Organism: Azoarcus olearius (bacteria)
SequenceString: GGCCGUGUGC CUUGCGCCGG GAAACCACGC AAGGGAUGGU GUCAAAUUCG GCGAAACCUA AGCGCGGAUG GUUCGCGUAU GGCAACGCCG AGCCAAGCUU CGCAGCCUUC GGGCUGCGAU GAAGGUGUAG AGACUAGACG GCACCCACCU AAGGCGCACC GAACCAUCCG ...String:
GGCCGUGUGC CUUGCGCCGG GAAACCACGC AAGGGAUGGU GUCAAAUUCG GCGAAACCUA AGCGCGGAUG GUUCGCGUAU GGCAACGCCG AGCCAAGCUU CGCAGCCUUC GGGCUGCGAU GAAGGUGUAG AGACUAGACG GCACCCACCU AAGGCGCACC GAACCAUCCG GUGCGCUAUG GUGAAGGCAU AGUCCAGGGA GUGGCGAAAG CCACACAAAC CAG

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Macromolecule #2: Ligated exon mimic of the Azoarcus Group I intron

MacromoleculeName: Ligated exon mimic of the Azoarcus Group I intron / type: other / ID: 2
Classification: polydeoxyribonucleotide/polyribonucleotide hybrid
Source (natural)Organism: Azoarcus olearius (bacteria)
SequenceString:
CATACGGCC

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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 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
20.0 mMTris-acetateTris-acetate
30.0 mMMgCl2Magnesium chloride
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 400
VitrificationCryogen name: ETHANE / Instrument: GATAN CRYOPLUNGE 3

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Electron microscopy

MicroscopeFEI POLARA 300
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 52.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: OTHER / Cs: 2.0 mm / Nominal magnification: 31000
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

Final reconstructionResolution.type: BY AUTHOR / Resolution: 4.9 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 486860
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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