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- EMDB-36777: The Anoxybacillus pushchinoensis ORF-less Group IIC Intron DR1 at... -

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Basic information

Entry
Database: EMDB / ID: EMD-36777
TitleThe Anoxybacillus pushchinoensis ORF-less Group IIC Intron DR1 at symmetric pre-cleavage state
Map data
Sample
  • Complex: The Anoxybacillus pushchinoensis ORF-less Group IIC Intron HYER1 at symmetric pre-cleavage state
    • Complex: RNA
      • RNA: RNA (542-MER)
    • Complex: DNA
      • DNA: DNA
  • Ligand: MAGNESIUM ION
KeywordsORF-less Group IIC intron / ribozymes / RNA
Biological speciesAnoxybacillus pushchinoensis (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.14 Å
AuthorsZhu HZ / Liu JJG
Funding support China, 1 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC) China
CitationJournal: Science / Year: 2024
Title: Hydrolytic endonucleolytic ribozyme (HYER) is programmable for sequence-specific DNA cleavage.
Authors: Zi-Xian Liu / Shouyue Zhang / Han-Zhou Zhu / Zhi-Hang Chen / Yun Yang / Long-Qi Li / Yuan Lei / Yun Liu / Dan-Yuan Li / Ao Sun / Cheng-Ping Li / Shun-Qing Tan / Gao-Li Wang / Jie-Yi Shen / ...Authors: Zi-Xian Liu / Shouyue Zhang / Han-Zhou Zhu / Zhi-Hang Chen / Yun Yang / Long-Qi Li / Yuan Lei / Yun Liu / Dan-Yuan Li / Ao Sun / Cheng-Ping Li / Shun-Qing Tan / Gao-Li Wang / Jie-Yi Shen / Shuai Jin / Caixia Gao / Jun-Jie Gogo Liu /
Abstract: Ribozymes are catalytic RNAs with diverse functions including self-splicing and polymerization. This work aims to discover natural ribozymes that behave as hydrolytic and sequence-specific DNA ...Ribozymes are catalytic RNAs with diverse functions including self-splicing and polymerization. This work aims to discover natural ribozymes that behave as hydrolytic and sequence-specific DNA endonucleases, which could be repurposed as DNA manipulation tools. Focused on bacterial group II-C introns, we found that many systems without intron-encoded protein propagate multiple copies in their resident genomes. These introns, named HYdrolytic Endonucleolytic Ribozymes (HYERs), cleaved RNA, single-stranded DNA, bubbled double-stranded DNA (dsDNA), and plasmids in vitro. HYER1 generated dsDNA breaks in the mammalian genome. Cryo-electron microscopy analysis revealed a homodimer structure for HYER1, where each monomer contains a Mg-dependent hydrolysis pocket and captures DNA complementary to the target recognition site (TRS). Rational designs including TRS extension, recruiting sequence insertion, and heterodimerization yielded engineered HYERs showing improved specificity and flexibility for DNA manipulation.
History
DepositionJul 10, 2023-
Header (metadata) releaseMay 15, 2024-
Map releaseMay 15, 2024-
UpdateMay 15, 2024-
Current statusMay 15, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_36777.png

Downloads & links

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Map

FileDownload / File: emd_36777.map.gz / Format: CCP4 / Size: 476.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
1.08 Å/pix.
x 500 pix.
= 541.25 Å		1.08 Å/pix.
x 500 pix.
= 541.25 Å		1.08 Å/pix.
x 500 pix.
= 541.25 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.0825 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.5
Minimum - Maximum-2.7684484 - 4.5840244
Average (Standard dev.)-0.00011256416 (±0.037964277)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions500500500
Spacing500500500
CellA=B=C: 541.25 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_36777_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_36777_half_map_2.map
Projections & Slices
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Histogram (log scale)

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Sample components

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Entire : The Anoxybacillus pushchinoensis ORF-less Group IIC Intron HYER1 ...

EntireName: The Anoxybacillus pushchinoensis ORF-less Group IIC Intron HYER1 at symmetric pre-cleavage state
Components
  • Complex: The Anoxybacillus pushchinoensis ORF-less Group IIC Intron HYER1 at symmetric pre-cleavage state
    • Complex: RNA
      • RNA: RNA (542-MER)
    • Complex: DNA
      • DNA: DNA
  • Ligand: MAGNESIUM ION

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Supramolecule #1: The Anoxybacillus pushchinoensis ORF-less Group IIC Intron HYER1 ...

SupramoleculeName: The Anoxybacillus pushchinoensis ORF-less Group IIC Intron HYER1 at symmetric pre-cleavage state
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Anoxybacillus pushchinoensis (bacteria)

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Supramolecule #2: RNA

SupramoleculeName: RNA / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1

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Supramolecule #3: DNA

SupramoleculeName: DNA / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2

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Macromolecule #1: RNA (542-MER)

MacromoleculeName: RNA (542-MER) / type: rna / ID: 1 / Number of copies: 2
Source (natural)Organism: Anoxybacillus pushchinoensis (bacteria)
Molecular weightTheoretical: 205.134922 KDa
SequenceString: GUGCGCUCGG CAUGGGUGCA AUCUCUAGGG UGAAAGUCCC GAACUGCGAA GGCAGAAGUA GCAGUUAGCU UAACGCAAGG GUGUCCGUG GUGACGCGGA AUCUGAAGGA AGCGGGCGGC AAACUUCCGG UCUGAGGAAC ACGAACUUCA UAUAAGGCUA G GUAUCAUU ...String:
GUGCGCUCGG CAUGGGUGCA AUCUCUAGGG UGAAAGUCCC GAACUGCGAA GGCAGAAGUA GCAGUUAGCU UAACGCAAGG GUGUCCGUG GUGACGCGGA AUCUGAAGGA AGCGGGCGGC AAACUUCCGG UCUGAGGAAC ACGAACUUCA UAUAAGGCUA G GUAUCAUU GGAUGAGUUU GCAAGACAAA ACAAAGUCCU UUCUGCCGAA GGUGAUACAG AGUAAAUGAA GCAGAUAGAU GG AAGGAAA GAUUGUACUC UUACCCGAGG AGGUCUGAUG GAUACGUGAA GUGCGCUUCA UAACCUACUU AGUGAUAAGU AAC UGAACC AUCAGAAGUC AGCAGAGGUC AUAGUACGAA UCGGUCUAGA ACGAUUCGGA AGGACUGAAC AAUCAAGAGA AAAU AGCCC UUGGCAUUCA GUACGUCAUG AUGAACACAG AAAACAUGGU ACCUCCCAAG AGAAAGGAAA CGGUGAAUCC CGUGG GAAU CUUUUGGAGG GUGGAGUGAC GACUGGCAUA AGAAGAUCAG CUAUUUACGG AAGGAAGCUU GCGUCAUUAU CUUGAU UGA ACCGCCGUAU ACGGAACCGU ACGUACGGUG GUGUGAGAGG ACGGAGGUUA AUCACCUCCU CCUACUCGAU

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Macromolecule #2: DNA

MacromoleculeName: DNA / type: dna / ID: 2 / Number of copies: 2 / Classification: DNA
Source (natural)Organism: Anoxybacillus pushchinoensis (bacteria)
Molecular weightTheoretical: 2.672758 KDa
SequenceString:
(DT)(DG)(DT)(DC)(DT)(DT)(DT)(DC)(DC)

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Macromolecule #3: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 3 / Number of copies: 4 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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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: 7.4
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / 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 / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.5 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.14 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 153769
Initial angle assignmentType: OTHER
Final angle assignmentType: OTHER

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