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- PDB-8xvd: CryoEM structure of ADP-DNA-MuB conformation2 -

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Basic information

Entry
Database: PDB / ID: 8xvd
TitleCryoEM structure of ADP-DNA-MuB conformation2
ComponentsATP-dependent target DNA activator B
KeywordsVIRAL PROTEIN / ADP-MuB complex / D10 symmetry / Ring
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
ADENOSINE-5'-DIPHOSPHATE / ATP-dependent target DNA activator B
Similarity search - Component
Biological speciesEscherichia phage Mu (virus)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.43 Å
AuthorsZhao, X. / Zhang, K. / Li, S.
Funding support1items
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, 2024Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Aug 14, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: ATP-dependent target DNA activator B
B: ATP-dependent target DNA activator B
C: ATP-dependent target DNA activator B
D: ATP-dependent target DNA activator B
E: ATP-dependent target DNA activator B
F: ATP-dependent target DNA activator B
G: ATP-dependent target DNA activator B
H: ATP-dependent target DNA activator B
I: ATP-dependent target DNA activator B
J: ATP-dependent target DNA activator B
K: ATP-dependent target DNA activator B
L: ATP-dependent target DNA activator B
M: ATP-dependent target DNA activator B
N: ATP-dependent target DNA activator B
O: ATP-dependent target DNA activator B
P: ATP-dependent target DNA activator B
Q: ATP-dependent target DNA activator B
R: ATP-dependent target DNA activator B
S: ATP-dependent target DNA activator B
T: ATP-dependent target DNA activator B
hetero molecules


Theoretical massNumber of molelcules
Total (without water)711,60740
Polymers703,06320
Non-polymers8,54420
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein
ATP-dependent target DNA activator B / Gene product 04 / gp04 / Gene product B / gpB / MuB


Mass: 35153.133 Da / Num. of mol.: 20
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia phage Mu (virus) / Gene: B, Mup04 / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21(DE3)
References: UniProt: P03763, Hydrolases; Acting on acid anhydrides; In phosphorus-containing anhydrides
#2: Chemical
ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE


Mass: 427.201 Da / Num. of mol.: 20 / Source method: obtained synthetically / Formula: C10H15N5O10P2 / Feature type: SUBJECT OF INVESTIGATION / Comment: ADP, energy-carrying molecule*YM
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: ADP-DNA-MuB conformation2 with D10 in Bacteriophage Mu DNA Transposition
Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Escherichia phage Mu (virus)
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria)
Buffer solutionpH: 7.8
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: BRIGHT FIELD / Nominal defocus max: 3000 nm / Nominal defocus min: 2000 nm
Image recordingElectron dose: 60.9 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of real images: 9510

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Processing

CTF correctionType: NONE
3D reconstructionResolution: 4.43 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 67932 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00438140
ELECTRON MICROSCOPYf_angle_d0.85351580
ELECTRON MICROSCOPYf_dihedral_angle_d4.73423460
ELECTRON MICROSCOPYf_chiral_restr0.0475940
ELECTRON MICROSCOPYf_plane_restr0.0066600

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