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- EMDB-38695: Cryo-EM structure of ATP-DNA-MuB filaments -

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Entry
Database: EMDB / ID: EMD-38695
TitleCryo-EM structure of ATP-DNA-MuB filaments
Map dataCryoEM structure of ATP-DNA-MuB Filaments in Bacteriophage Mu DNA Transposition
Sample
  • Complex: ATP-DNA-MuB Filaments in Bacteriophage Mu DNA Transposition
    • DNA: DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')
    • Protein or peptide: ATP-dependent target DNA activator B
    • DNA: DNA (5'-D(P*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*T)-3')
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
KeywordsVIRAL PROTEIN-DNA COMPLEX
Function / homology
Function and homology information


DNA transposition / viral DNA genome replication / Hydrolases; Acting on acid anhydrides; In phosphorus-containing anhydrides / DNA integration / DNA replication / host cell cytoplasm / ATP hydrolysis activity / DNA binding / ATP binding / metal ion binding
Similarity search - Function
B transposition protein, C-terminal / B transposition protein, C-terminal domain superfamily / Mu B transposition protein, C terminal / : / : / AAA domain / Lambda repressor-like, DNA-binding domain superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
ATP-dependent target DNA activator B
Similarity search - Component
Biological speciesEscherichia phage Mu (virus)
Methodhelical reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsZhao X / Zhang K / Li S
Funding support1 items
OrganizationGrant numberCountry
Other government
CitationJournal: Nat Commun / Year: 2024
Title: Elucidating the Architectural dynamics of MuB filaments in bacteriophage Mu DNA transposition.
Authors: Xiaolong Zhao / Yongxiang Gao / Qingguo Gong / Kaiming Zhang / Shanshan Li /
Abstract: MuB is a non-specific DNA-binding protein and AAA+ ATPase that significantly influences the DNA transposition process of bacteriophage Mu, especially in target DNA selection for transposition. While ...MuB is a non-specific DNA-binding protein and AAA+ ATPase that significantly influences the DNA transposition process of bacteriophage Mu, especially in target DNA selection for transposition. While studies have established the ATP-dependent formation of MuB filament as pivotal to this process, the high-resolution structure of a full-length MuB protomer and the underlying molecular mechanisms governing its oligomerization remain elusive. Here, we use cryo-EM to obtain a 3.4-Å resolution structure of the ATP(+)-DNA(+)-MuB helical filament, which encapsulates the DNA substrate within its axial channel. The structure categorizes MuB within the initiator clade of the AAA+ protein family and precisely locates the ATP and DNA binding sites. Further investigation into the oligomeric states of MuB show the existence of various forms of the filament. These findings lead to a mechanistic model where MuB forms opposite helical filaments along the DNA, exposing potential target sites on the bare DNA and then recruiting MuA, which stimulates MuB's ATPase activity and disrupts the previously formed helical structure. When this happens, MuB generates larger ring structures and dissociates from the DNA.
History
DepositionJan 14, 2024-
Header (metadata) releaseAug 14, 2024-
Map releaseAug 14, 2024-
UpdateAug 14, 2024-
Current statusAug 14, 2024Processing site: PDBj / Status: Released

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Structure visualization

Downloads & links

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Map

FileDownload / File: emd_38695.map.gz / Format: CCP4 / Size: 144.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryoEM structure of ATP-DNA-MuB Filaments in Bacteriophage Mu DNA Transposition
Voxel sizeX=Y=Z: 0.82 Å
Density
Contour LevelBy AUTHOR: 0.1
Minimum - Maximum-0.073785104 - 0.23704728
Average (Standard dev.)0.004625997 (±0.015883258)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions336336336
Spacing336336336
CellA=B=C: 275.52 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : ATP-DNA-MuB Filaments in Bacteriophage Mu DNA Transposition

EntireName: ATP-DNA-MuB Filaments in Bacteriophage Mu DNA Transposition
Components
  • Complex: ATP-DNA-MuB Filaments in Bacteriophage Mu DNA Transposition
    • DNA: DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')
    • Protein or peptide: ATP-dependent target DNA activator B
    • DNA: DNA (5'-D(P*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*T)-3')
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE

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Supramolecule #1: ATP-DNA-MuB Filaments in Bacteriophage Mu DNA Transposition

SupramoleculeName: ATP-DNA-MuB Filaments in Bacteriophage Mu DNA Transposition
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3
Source (natural)Organism: Escherichia phage Mu (virus)

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Macromolecule #1: DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP...

MacromoleculeName: DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')
type: dna / ID: 1 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Escherichia phage Mu (virus)
Molecular weightTheoretical: 7.472007 KDa
SequenceString:
(DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA) (DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA) (DA)(DA)(DA)(DA)

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Macromolecule #3: DNA (5'-D(P*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP...

MacromoleculeName: DNA (5'-D(P*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*T)-3')
type: dna / ID: 3 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Escherichia phage Mu (virus)
Molecular weightTheoretical: 7.25567 KDa
SequenceString:
(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)

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Macromolecule #2: ATP-dependent target DNA activator B

MacromoleculeName: ATP-dependent target DNA activator B / type: protein_or_peptide / ID: 2 / Number of copies: 8 / Enantiomer: LEVO
EC number: Hydrolases; Acting on acid anhydrides; In phosphorus-containing anhydrides
Source (natural)Organism: Escherichia phage Mu (virus)
Molecular weightTheoretical: 35.153133 KDa
Recombinant expressionOrganism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
SequenceString: MNISDIRAGL RTLVENEETT FKQIALESGL STGTISSFIN DKYNGDNERV SQMLQRWLEK YHAVAELPEP PRFVETQTVK QIWTSMRFA SLTESIAVVC GNPGVGKTEA AREYRRTNNN VWMITITPSC ASVLECLTEL AFELGMNDAP RRKGPLSRAL R RRLEGTQG ...String:
MNISDIRAGL RTLVENEETT FKQIALESGL STGTISSFIN DKYNGDNERV SQMLQRWLEK YHAVAELPEP PRFVETQTVK QIWTSMRFA SLTESIAVVC GNPGVGKTEA AREYRRTNNN VWMITITPSC ASVLECLTEL AFELGMNDAP RRKGPLSRAL R RRLEGTQG LVIIDEADHL GAEVLEELRL LQESTRIGLV LMGNHRVYSN MTGGNRTVEF ARLFSRIAKR TAINKTKKAD VK AIADAWQ INGEKELELL QQIAQKPGAL RILNHSLRLA AMTAHGKGER VNEDYLRQAF RELDLDVDIS TLLRN

UniProtKB: ATP-dependent target DNA activator B

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Macromolecule #4: ADENOSINE-5'-TRIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 4 / Number of copies: 8 / Formula: ATP
Molecular weightTheoretical: 507.181 Da
Chemical component information

ChemComp-ATP:
ADENOSINE-5'-TRIPHOSPHATE / ATP, energy-carrying molecule*YM

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statefilament

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Sample preparation

BufferpH: 7.8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number real images: 6298 / Average electron dose: 60.9 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.0 µm / Nominal defocus min: 2.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Helical parameters - Δz: 6.91 Å
Applied symmetry - Helical parameters - Δ&Phi: 56.16 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Resolution.type: BY AUTHOR / Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 377689
Startup modelType of model: NONE
Final angle assignmentType: NOT APPLICABLE

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