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- PDB-4bt0: MuB is an AAAplus ATPase that forms helical filaments to control ... -

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Basic information

Entry
Database: PDB / ID: 4bt0
TitleMuB is an AAAplus ATPase that forms helical filaments to control target selection for DNA transposition
Components(TRANSCRIPTIONAL REGULATOR) x 2
KeywordsTRANSCRIPTION / AAA+ ATPASE / DNA TRANSPOSITION / NUCLEOPROTEIN FILAMENT / SYMMETRY MISMATCH
Function / homology
Function and homology information


phosphorelay signal transduction system / sequence-specific DNA binding / regulation of DNA-templated transcription / ATP hydrolysis activity / ATP binding / identical protein binding / metal ion binding
Similarity search - Function
Sigma-54 interaction domain ATP-binding region A signature. / Sigma-54 interaction domain, ATP-binding site 2 / Sigma-54 interaction domain ATP-binding region B signature. / Sigma-54 interaction domain, ATP-binding site 1 / Sigma-54 interaction domain profile. / Sigma-54 interaction domain / RNA polymerase sigma factor 54 interaction domain / DNA binding HTH domain, Fis-type / Bacterial regulatory protein, Fis family / Response regulator receiver domain ...Sigma-54 interaction domain ATP-binding region A signature. / Sigma-54 interaction domain, ATP-binding site 2 / Sigma-54 interaction domain ATP-binding region B signature. / Sigma-54 interaction domain, ATP-binding site 1 / Sigma-54 interaction domain profile. / Sigma-54 interaction domain / RNA polymerase sigma factor 54 interaction domain / DNA binding HTH domain, Fis-type / Bacterial regulatory protein, Fis family / Response regulator receiver domain / cheY-homologous receiver domain / Signal transduction response regulator, receiver domain / Response regulatory domain profile. / CheY-like superfamily / Homeobox-like domain superfamily / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
ADENOSINE-5'-DIPHOSPHATE / Transcriptional regulator (NtrC family)
Similarity search - Component
Biological speciesENTEROBACTERIA PHAGE MU (virus)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 17 Å
AuthorsMizuno, N. / Dramicanin, M. / Mizuuchi, M. / Adam, J. / Wang, Y. / Han, Y.W. / Yang, W. / Steven, A.C. / Mizuuchi, K. / Ramon-Maiques, S.
CitationJournal: Proc Natl Acad Sci U S A / Year: 2013
Title: MuB is an AAA+ ATPase that forms helical filaments to control target selection for DNA transposition.
Authors: Naoko Mizuno / Marija Dramićanin / Michiyo Mizuuchi / Julia Adam / Yi Wang / Yong-Woon Han / Wei Yang / Alasdair C Steven / Kiyoshi Mizuuchi / Santiago Ramón-Maiques /
Abstract: MuB is an ATP-dependent nonspecific DNA-binding protein that regulates the activity of the MuA transposase and captures target DNA for transposition. Mechanistic understanding of MuB function has ...MuB is an ATP-dependent nonspecific DNA-binding protein that regulates the activity of the MuA transposase and captures target DNA for transposition. Mechanistic understanding of MuB function has previously been hindered by MuB's poor solubility. Here we combine bioinformatic, mutagenic, biochemical, and electron microscopic analyses to unmask the structure and function of MuB. We demonstrate that MuB is an ATPase associated with diverse cellular activities (AAA+ ATPase) and forms ATP-dependent filaments with or without DNA. We also identify critical residues for MuB's ATPase, DNA binding, protein polymerization, and MuA interaction activities. Using single-particle electron microscopy, we show that MuB assembles into a helical filament, which binds the DNA in the axial channel. The helical parameters of the MuB filament do not match those of the coated DNA. Despite this protein-DNA symmetry mismatch, MuB does not deform the DNA duplex. These findings, together with the influence of MuB filament size on strand-transfer efficiency, lead to a model in which MuB-imposed symmetry transiently deforms the DNA at the boundary of the MuB filament and results in a bent DNA favored by MuA for transposition.
History
DepositionJun 12, 2013Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jul 3, 2013Provider: repository / Type: Initial release
Revision 1.1Jul 17, 2013Group: Database references
Revision 1.2Aug 7, 2013Group: Derived calculations
Revision 1.3Aug 23, 2017Group: Data collection / Category: em_software
Item: _em_software.fitting_id / _em_software.image_processing_id
Revision 1.4May 8, 2024Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / em_3d_fitting_list / pdbx_initial_refinement_model / pdbx_struct_oper_list / struct_site
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type / _pdbx_struct_oper_list.name / _pdbx_struct_oper_list.symmetry_operation / _pdbx_struct_oper_list.type / _struct_site.pdbx_auth_asym_id / _struct_site.pdbx_auth_comp_id / _struct_site.pdbx_auth_seq_id

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Structure visualization

Movie
  • Biological unit as representative helical assembly
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  • Deposited structure unit
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  • Simplified surface model + fitted atomic model
  • EMDB-2398
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  • Superimposition on EM map
  • EMDB-2398
  • Imaged by UCSF Chimera
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Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: TRANSCRIPTIONAL REGULATOR
B: TRANSCRIPTIONAL REGULATOR
hetero molecules


Theoretical massNumber of molelcules
Total (without water)28,8154
Polymers27,9602
Non-polymers8542
Water00
1
A: TRANSCRIPTIONAL REGULATOR
B: TRANSCRIPTIONAL REGULATOR
hetero molecules
x 7


Theoretical massNumber of molelcules
Total (without water)201,70328
Polymers195,72214
Non-polymers5,98114
Water0
TypeNameSymmetry operationNumber
helical symmetry operation6
identity operation1_555x,y,z1
2


  • Idetical with deposited unit
  • helical asymmetric unit
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
3


  • Idetical with deposited unit
  • helical asymmetric unit, std helical frame
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
SymmetryHelical symmetry: (Circular symmetry: 1 / Dyad axis: no / N subunits divisor: 1 / Num. of operations: 7 / Rise per n subunits: 9.01 Å / Rotation per n subunits: 66.2 °)
Noncrystallographic symmetry (NCS)NCS oper:
IDCodeMatrixVector
1generate(-0.947768, -0.318959), (0.318959, -0.947768), (1)-27.03
2generate(-0.674302, 0.738455), (-0.738455, -0.674302), (1)-18.02
3generate(0.403545, 0.91496), (-0.91496, 0.403545), (1)-9.01
4given(1), (1), (1)
5generate(0.403545, -0.91496), (0.91496, 0.403545), (1)9.01
6generate(-0.674302, -0.738455), (0.738455, -0.674302), (1)18.02
7generate(-0.947768, 0.318959), (-0.318959, -0.947768), (1)27.03

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Components

#1: Protein TRANSCRIPTIONAL REGULATOR / MUB AAAPLUS ATPASE


Mass: 8562.894 Da / Num. of mol.: 1 / Fragment: AAAPLUS DOMAIN, RESIDUES 312-384
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) ENTEROBACTERIA PHAGE MU (virus) / Production host: ESCHERICHIA COLI (E. coli) / References: UniProt: O67198
#2: Protein TRANSCRIPTIONAL REGULATOR / MUB AAAPLUS ATPASE


Mass: 19397.400 Da / Num. of mol.: 1 / Fragment: AAAPLUS DOMAIN, RESIDUES 137-309
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) ENTEROBACTERIA PHAGE MU (virus) / Production host: ESCHERICHIA COLI (E. coli) / References: UniProt: O67198
#3: Chemical ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE


Mass: 427.201 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C10H15N5O10P2 / Comment: ADP, energy-carrying molecule*YM

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Experimental details

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Experiment

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

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Sample preparation

ComponentName: MUB FILAMENT WITH DNA / Type: COMPLEX
Buffer solutionName: 30 MM TRISHCL PH 8.0, 0.3 M KCL, 5MM MGCL2, 1MM DTT, 1 MM ATP OR ATP-GAMMA-S
pH: 8
Details: 30 MM TRISHCL PH 8.0, 0.3 M KCL, 5MM MGCL2, 1MM DTT, 1 MM ATP OR ATP-GAMMA-S
SpecimenConc.: 0.07 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: HOLEY CARBON
VitrificationInstrument: FEI VITROBOT MARK II / Cryogen name: ETHANE / Details: LIQUID ETHANE

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Electron microscopy imaging

MicroscopyModel: FEI/PHILIPS CM200FEG / Date: Apr 27, 2008
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 120 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 38000 X / Nominal defocus max: 3000 nm / Nominal defocus min: 1500 nm
Specimen holderTemperature: 82 K
Image recordingElectron dose: 15 e/Å2 / Film or detector model: GATAN ULTRASCAN 1000 (2k x 2k)
Image scansNum. digital images: 109

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Processing

EM software
IDNameCategory
1UCSF Chimeramodel fitting
2Bsoft3D reconstruction
3EMAN3D reconstruction
4SPIDER3D reconstruction
CTF correctionDetails: PHASE-FLIPPING
3D reconstructionMethod: IHRSR / Resolution: 17 Å / Nominal pixel size: 2.8 Å / Actual pixel size: 2.8 Å
Magnification calibration: SUBMISSION BASED ON EXPERIMENTAL DATA FROM EMDB EMD-2398. (DEPOSITION ID: 11698)
Symmetry type: HELICAL
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Details: METHOD--RIGID BODY REFINEMENT PROTOCOL--ELECTRON MICROSCPY
Atomic model buildingPDB-ID: 1NY6

1ny6


Accession code: 1NY6 / Source name: PDB / Type: experimental model
RefinementHighest resolution: 17 Å
Refinement stepCycle: LAST / Highest resolution: 17 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms1969 0 54 0 2023

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