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- PDB-8t2r: Structure of a group II intron ribonucleoprotein in the pre-ligat... -

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Basic information

Entry
Database: PDB / ID: 8t2r
TitleStructure of a group II intron ribonucleoprotein in the pre-ligation (pre-2F) state
Components
  • 5'exon
  • Group II intron reverse transcriptase/maturase
  • Intron
KeywordsTransferase/RNA / RNP / RNA / Transferase-RNA complex
Function / homology
Function and homology information


RNA-directed DNA polymerase / RNA-directed DNA polymerase activity
Similarity search - Function
Group II intron, maturase-specific / Group II intron reverse transcriptase/maturase / Group II intron, maturase-specific domain / : / Reverse transcriptase domain / Reverse transcriptase (RT) catalytic domain profile. / Reverse transcriptase (RNA-dependent DNA polymerase) / DNA/RNA polymerase superfamily
Similarity search - Domain/homology
AMMONIUM ION / 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: 3.1 Å
AuthorsXu, L. / Liu, T. / Chung, K. / Pyle, A.M.
Funding support United States, 1items
OrganizationGrant numberCountry
Howard Hughes Medical Institute (HHMI) United States
CitationJournal: Nature / Year: 2023
Title: Structural insights into intron catalysis and dynamics during splicing.
Authors: Ling Xu / Tianshuo Liu / Kevin Chung / Anna Marie Pyle /
Abstract: The group II intron ribonucleoprotein is an archetypal splicing system with numerous mechanistic parallels to the spliceosome, including excision of lariat introns. Despite the importance of ...The group II intron ribonucleoprotein is an archetypal splicing system with numerous mechanistic parallels to the spliceosome, including excision of lariat introns. Despite the importance of branching in RNA metabolism, structural understanding of this process has remained elusive. Here we present a comprehensive analysis of three single-particle cryogenic electron microscopy structures captured along the splicing pathway. They reveal the network of molecular interactions that specifies the branchpoint adenosine and positions key functional groups to catalyse lariat formation and coordinate exon ligation. The structures also reveal conformational rearrangements of the branch helix and the mechanism of splice site exchange that facilitate the transition from branching to ligation. These findings shed light on the evolution of splicing and highlight the conservation of structural components, catalytic mechanism and dynamical strategies retained through time in premessenger RNA splicing machines.
History
DepositionJun 6, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Nov 22, 2023Provider: repository / Type: Initial release
Revision 1.1Dec 6, 2023Group: Database references / Category: citation / Item: _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.2Jan 3, 2024Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
D: Group II intron reverse transcriptase/maturase
A: 5'exon
B: Intron
hetero molecules


Theoretical massNumber of molelcules
Total (without water)260,13418
Polymers259,5773
Non-polymers55715
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: 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
#2: RNA chain 5'exon


Mass: 2097.219 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) [Eubacterium] rectale (bacteria) / Production host: Escherichia coli (E. coli)
#3: RNA chain Intron


Mass: 208395.812 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) [Eubacterium] rectale (bacteria) / Production host: Escherichia coli (E. coli)
#4: Chemical
ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 13 / Source method: obtained synthetically / Formula: Ca / Feature type: SUBJECT OF INVESTIGATION
#5: Chemical ChemComp-NH4 / AMMONIUM ION


Mass: 18.038 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: H4N / Feature type: SUBJECT OF INVESTIGATION
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: Intron RNP complex in pre-2f / Type: COMPLEX / Entity ID: #1-#3 / Source: RECOMBINANT
Molecular weightValue: 0.3 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
VitrificationCryogen name: ETHANE

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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: DARK FIELD / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm
Image recordingElectron dose: 1.5 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM softwareName: PHENIX / Version: 1.20.1_4487: / Category: model refinement
CTF correctionType: NONE
3D reconstructionResolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 234625 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00616534
ELECTRON MICROSCOPYf_angle_d0.94825072
ELECTRON MICROSCOPYf_dihedral_angle_d18.3547043
ELECTRON MICROSCOPYf_chiral_restr0.0453232
ELECTRON MICROSCOPYf_plane_restr0.0071111

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