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- PDB-7v9x: Cryo-EM structure of E.coli retron-Ec86 in complex with its effec... -

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Basic information

Entry
Database: PDB / ID: 7v9x
TitleCryo-EM structure of E.coli retron-Ec86 in complex with its effector at 2.8 angstrom
Components
  • DNA (105-MER)
  • RNA (14-MER)
  • RNA (81-MER)
  • RNA-directed DNA polymerase from retron EC86
  • retron St85 family effector protein
KeywordsTRANSFERASE/RNA BINDING PROTEIN/DNA/RNA / Reverse transcriptase / RNA BINDING PROTEIN / TRANSFERASE-RNA BINDING PROTEIN-DNA-RNA complex
Function / homology
Function and homology information


RNA-directed DNA polymerase / RNA-directed DNA polymerase activity / defense response to virus / RNA binding / metal ion binding
Similarity search - Function
RNA-directed DNA polymerase (reverse transcriptase), msDNA / Reverse transcriptase (RNA-dependent DNA polymerase) / Reverse transcriptase domain / Reverse transcriptase (RT) catalytic domain profile. / DNA/RNA polymerase superfamily
Similarity search - Domain/homology
: / DNA / DNA (> 10) / DNA (> 100) / RNA / RNA (> 10) / : / Retron Ec86 reverse transcriptase
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.82 Å
AuthorsWang, Y.J. / Guan, Z.Y. / Zou, T.T.
Funding support China, 2items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)2018YFA0507700 China
National Natural Science Foundation of China (NSFC)32070174 China
CitationJournal: To Be Published
Title: Structural insight into anti-phage Retron-Ec86 complex
Authors: Wang, T.J. / Guan, Z.Y. / Zou, T.T.
History
DepositionAug 27, 2021Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Aug 31, 2022Provider: repository / Type: Initial release
Revision 1.1Jun 19, 2024Group: Data collection / Category: chem_comp_atom / chem_comp_bond

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: RNA-directed DNA polymerase from retron EC86
C: retron St85 family effector protein
D: DNA (105-MER)
E: RNA (81-MER)
F: RNA (14-MER)
B: RNA-directed DNA polymerase from retron EC86
G: DNA (105-MER)
H: RNA (81-MER)
I: RNA (14-MER)


Theoretical massNumber of molelcules
Total (without water)236,9649
Polymers236,9649
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area29380 Å2
ΔGint-84 kcal/mol
Surface area61630 Å2

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Components

#1: Protein RNA-directed DNA polymerase from retron EC86 / EC86-RT / Reverse transcriptase


Mass: 37832.070 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P23070, RNA-directed DNA polymerase
#2: Protein retron St85 family effector protein


Mass: 35538.242 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Gene: HJV81_000169 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: A0A7A0NNW9
#3: DNA chain DNA (105-MER)


Mass: 32468.803 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Production host: Escherichia coli BL21(DE3) (bacteria) / References: GenBank: 341167
#4: RNA chain RNA (81-MER)


Mass: 25870.203 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Production host: Escherichia coli BL21(DE3) (bacteria) / References: GenBank: 341167
#5: RNA chain RNA (14-MER)


Mass: 4541.771 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Production host: Escherichia coli BL21(DE3) (bacteria)

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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: retron-Ec86 (RT-msDNA-RNA) / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Escherichia coli (E. coli)
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria)
Buffer solutionpH: 8
SpecimenConc.: 0.4 mg/ml / Embedding 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: BRIGHT FIELD
Image recordingElectron dose: 53.68 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.19.1_4122: / Classification: refinement
EM softwareName: cryoSPARC / Version: 3.1 / Category: final Euler assignment
CTF correctionType: NONE
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 2.82 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 137409 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00511190
ELECTRON MICROSCOPYf_angle_d0.61816051
ELECTRON MICROSCOPYf_dihedral_angle_d23.1753181
ELECTRON MICROSCOPYf_chiral_restr0.0391916
ELECTRON MICROSCOPYf_plane_restr0.0041259

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