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- PDB-7uin: CryoEM Structure of an Group II Intron Retroelement -

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Basic information

Entry
Database: PDB / ID: 7uin
TitleCryoEM Structure of an Group II Intron Retroelement
Components
  • DNA (37-MER)
  • E.r IIC Intron
  • Group II intron reverse transcriptase/maturase
KeywordsRNA BINDING PROTEIN/RNA/DNA / RNA / intron / group II / maturase / splicing / retrotransposition / RNA BINDING PROTEIN-RNA-DNA complex
Function / homology
Function and homology information


RNA-directed DNA polymerase / RNA-directed DNA polymerase activity / DNA damage response
Similarity search - Function
Group II intron, maturase-specific / Group II intron reverse transcriptase/maturase / Group II intron, maturase-specific domain / Reverse transcriptase (RNA-dependent DNA polymerase) / Reverse transcriptase domain / Reverse transcriptase (RT) catalytic domain profile. / DNA/RNA polymerase superfamily
Similarity search - Domain/homology
AMMONIUM ION / DNA / DNA (> 10) / RNA / RNA (> 10) / RNA (> 100) / Group II intron reverse transcriptase/maturase
Similarity search - Component
Biological species[Eubacterium] rectale (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.8 Å
AuthorsChung, K. / Xu, L.
Funding support United States, 1items
OrganizationGrant numberCountry
Howard Hughes Medical Institute (HHMI) United States
CitationJournal: Science / Year: 2022
Title: Structures of a mobile intron retroelement poised to attack its structured DNA target.
Authors: Kevin Chung / Ling Xu / Pengxin Chai / Junhui Peng / Swapnil C Devarkar / Anna Marie Pyle /
Abstract: Group II introns are ribozymes that catalyze their self-excision and function as retroelements that invade DNA. As retrotransposons, group II introns form ribonucleoprotein (RNP) complexes that roam ...Group II introns are ribozymes that catalyze their self-excision and function as retroelements that invade DNA. As retrotransposons, group II introns form ribonucleoprotein (RNP) complexes that roam the genome, integrating by reversal of forward splicing. Here we show that retrotransposition is achieved by a tertiary complex between a structurally elaborate ribozyme, its protein mobility factor, and a structured DNA substrate. We solved cryo-electron microscopy structures of an intact group IIC intron-maturase retroelement that was poised for integration into a DNA stem-loop motif. By visualizing the RNP before and after DNA targeting, we show that it is primed for attack and fits perfectly with its DNA target. This study reveals design principles of a prototypical retroelement and reinforces the hypothesis that group II introns are ancient elements of genetic diversification.
History
DepositionMar 29, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Nov 23, 2022Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
B: E.r IIC Intron
D: Group II intron reverse transcriptase/maturase
A: DNA (37-MER)
hetero molecules


Theoretical massNumber of molelcules
Total (without water)267,98433
Polymers267,2673
Non-polymers71730
Water43224
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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RNA chain / Protein / DNA chain , 3 types, 3 molecules BDA

#1: RNA chain E.r IIC Intron


Mass: 206779.859 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) [Eubacterium] rectale (bacteria)
#2: Protein Group II intron reverse transcriptase/maturase / Group II intron-encoded protein ltrA


Mass: 49083.914 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) [Eubacterium] rectale (bacteria) / Gene: ltrA_2, ltrA, ERS852417_00966, FYL37_05080 / Production host: Escherichia coli (E. coli)
References: UniProt: A0A173ZME3, RNA-directed DNA polymerase
#3: DNA chain DNA (37-MER)


Mass: 11403.316 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) [Eubacterium] rectale (bacteria)

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Non-polymers , 3 types, 54 molecules

#4: Chemical...
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 28 / Source method: obtained synthetically / Formula: Mg / Feature type: SUBJECT OF INVESTIGATION
#5: Chemical ChemComp-NH4 / AMMONIUM ION / Ammonium


Mass: 18.038 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: H4N / Feature type: SUBJECT OF INVESTIGATION
#6: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 24 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Ternary complex of RNA, protein and DNA / Type: COMPLEX / Entity ID: #1-#3 / Source: RECOMBINANT
Molecular weightValue: 0.260 MDa / Experimental value: NO
Source (natural)Organism: [Eubacterium] rectale (bacteria)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 297 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2500 nm / Nominal defocus min: 500 nm
Specimen holderCryogen: NITROGEN
Image recordingElectron dose: 50.5 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM software
IDNameVersionCategory
2SerialEMimage acquisition
4cryoSPARC3.1CTF correction
9cryoSPARC3.1initial Euler assignment
10cryoSPARC3.1final Euler assignment
12cryoSPARC3.13D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 914099 / Symmetry type: POINT

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