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- EMDB-2333: Structural insights into the chaperone activity of Hsp40: DnaJ bi... -

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Basic information

Entry
Database: EMDB / ID: EMD-2333
TitleStructural insights into the chaperone activity of Hsp40: DnaJ binds and remodels RepE
Map data3D Resonstruction of DnaJ:RepE(1-144) complex
Sample
  • Sample: DnaJ:RepE(1-144) complex
  • Protein or peptide: Hsp40
  • Protein or peptide: Shorter version of RepE (RepE 1-144)
KeywordsHsp40 / DnaJ / RepE / chaperones / protein folding / electron microscopy
Function / homology
Function and homology information


sigma factor antagonist activity / protein disulfide isomerase activity / chaperone cofactor-dependent protein refolding / protein unfolding / protein-disulfide reductase activity / viral process / heat shock protein binding / unfolded protein binding / protein folding / protein-folding chaperone binding ...sigma factor antagonist activity / protein disulfide isomerase activity / chaperone cofactor-dependent protein refolding / protein unfolding / protein-disulfide reductase activity / viral process / heat shock protein binding / unfolded protein binding / protein folding / protein-folding chaperone binding / response to heat / protein refolding / protein-containing complex assembly / DNA replication / protein homodimerization activity / protein-containing complex / zinc ion binding / ATP binding / membrane / cytosol / cytoplasm
Similarity search - Function
Chaperone DnaJ / DnaJ central domain / Heat shock protein DnaJ, cysteine-rich domain / Zinc finger CR-type profile. / Heat shock protein DnaJ, cysteine-rich domain superfamily / HSP40/DnaJ peptide-binding / Chaperone DnaJ, C-terminal / DnaJ C terminal domain / Nt-dnaJ domain signature. / DnaJ domain, conserved site ...Chaperone DnaJ / DnaJ central domain / Heat shock protein DnaJ, cysteine-rich domain / Zinc finger CR-type profile. / Heat shock protein DnaJ, cysteine-rich domain superfamily / HSP40/DnaJ peptide-binding / Chaperone DnaJ, C-terminal / DnaJ C terminal domain / Nt-dnaJ domain signature. / DnaJ domain, conserved site / DnaJ domain / DnaJ molecular chaperone homology domain / dnaJ domain profile. / Chaperone J-domain superfamily / DnaJ domain
Similarity search - Domain/homology
Chaperone protein DnaJ
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / negative staining / Resolution: 20.0 Å
AuthorsCuellar J / Perales-Calvo J / Muga A / Valpuesta JM / Moro F
CitationJournal: J Biol Chem / Year: 2013
Title: Structural insights into the chaperone activity of the 40-kDa heat shock protein DnaJ: binding and remodeling of a native substrate.
Authors: Jorge Cuéllar / Judit Perales-Calvo / Arturo Muga / José María Valpuesta / Fernando Moro /
Abstract: Hsp40 chaperones bind and transfer substrate proteins to Hsp70s and regulate their ATPase activity. The interaction of Hsp40s with native proteins modifies their structure and function. A good model ...Hsp40 chaperones bind and transfer substrate proteins to Hsp70s and regulate their ATPase activity. The interaction of Hsp40s with native proteins modifies their structure and function. A good model for this function is DnaJ, the bacterial Hsp40 that interacts with RepE, the repressor/activator of plasmid F replication, and together with DnaK regulates its function. We characterize here the structure of the DnaJ-RepE complex by electron microscopy, the first described structure of a complex between an Hsp40 and a client protein. The comparison of the complexes of DnaJ with two RepE mutants reveals an intrinsic plasticity of the DnaJ dimer that allows the chaperone to adapt to different substrates. We also show that DnaJ induces conformational changes in dimeric RepE, which increase the intermonomeric distance and remodel both RepE domains enhancing its affinity for DNA.
History
DepositionMar 15, 2013-
Header (metadata) releaseApr 17, 2013-
Map releaseApr 24, 2013-
UpdateDec 25, 2013-
Current statusDec 25, 2013Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 2.7
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 2.7
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd2333.jpg

Downloads & links

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Map

FileDownload / File: emd_2333.map.gz / Format: CCP4 / Size: 1.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation3D Resonstruction of DnaJ:RepE(1-144) complex
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
2.33 Å/pix.
x 80 pix.
= 186.4 Å		2.33 Å/pix.
x 80 pix.
= 186.4 Å		2.33 Å/pix.
x 80 pix.
= 186.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 2.33 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 2.7 / Movie #1: 2.7
Minimum - Maximum-3.54815888 - 10.211532589999999
Average (Standard dev.)0.0 (±0.99999994)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-40-40-40
Dimensions808080
Spacing808080
CellA=B=C: 186.4 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.332.332.33
M x/y/z808080
origin x/y/z0.0000.0000.000
length x/y/z186.400186.400186.400
α/β/γ90.00090.00090.000
start NX/NY/NZ-27-15-36
NX/NY/NZ553173
MAP C/R/S123
start NC/NR/NS-40-40-40
NC/NR/NS808080
D min/max/mean-3.54810.212-0.000

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Supplemental data

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Sample components

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Entire : DnaJ:RepE(1-144) complex

EntireName: DnaJ:RepE(1-144) complex
Components
  • Sample: DnaJ:RepE(1-144) complex
  • Protein or peptide: Hsp40
  • Protein or peptide: Shorter version of RepE (RepE 1-144)

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Supramolecule #1000: DnaJ:RepE(1-144) complex

SupramoleculeName: DnaJ:RepE(1-144) complex / type: sample / ID: 1000
Oligomeric state: One dimer of DnaJ binds to one dimer of RepE
Number unique components: 2
Molecular weightExperimental: 115 KDa / Theoretical: 115 KDa / Method: Analytical ultracentrifugation (AU)

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Macromolecule #1: Hsp40

MacromoleculeName: Hsp40 / type: protein_or_peptide / ID: 1 / Name.synonym: DnaJ / Number of copies: 2 / Oligomeric state: dimer / Recombinant expression: Yes
Source (natural)Organism: Escherichia coli (E. coli) / Location in cell: cytoplasm
Molecular weightExperimental: 80 KDa / Theoretical: 80 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceUniProtKB: Chaperone protein DnaJ

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Macromolecule #2: Shorter version of RepE (RepE 1-144)

MacromoleculeName: Shorter version of RepE (RepE 1-144) / type: protein_or_peptide / ID: 2 / Name.synonym: RepE(1-154) / Number of copies: 2 / Oligomeric state: dimer / Recombinant expression: Yes
Source (natural)Organism: Escherichia coli (E. coli) / Location in cell: cytoplasm
Molecular weightExperimental: 32 KDa / Theoretical: 32 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)

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Experimental details

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Structure determination

Methodnegative staining
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.4 mg/mL
BufferpH: 7.4
Details: 20mM Hepes pH 7.4, 50mM KCl, 5mM DTT and 0.1mM EDTA
StainingType: NEGATIVE
Details: Samples (either DnaJ:RepE, DnaJ:RepE1-144 or DnaJ:RepE54 complexes) were diluted 1:100 in 20mM Hepes pH 7.4, 50mM KCl, 5mM DTT and 0.1mM EDTA buffer, applied onto carbon-coated copper grids ...Details: Samples (either DnaJ:RepE, DnaJ:RepE1-144 or DnaJ:RepE54 complexes) were diluted 1:100 in 20mM Hepes pH 7.4, 50mM KCl, 5mM DTT and 0.1mM EDTA buffer, applied onto carbon-coated copper grids and stained with 2% uranyl acetate.
GridDetails: 300 mesh Cu/Rh grid with thin carbon support and glow discharged
VitrificationCryogen name: NONE / Instrument: OTHER

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Electron microscopy

MicroscopeJEOL 1200EXII
Alignment procedureLegacy - Astigmatism: Objective lens astigmatism was corrected at 100,000 times magnification
DetailsAdditional details about microscope model:JEOL 1200EXII
DateJun 8, 2011
Image recordingCategory: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: ZEISS SCAI / Digitization - Sampling interval: 14 µm / Number real images: 270 / Average electron dose: 10 e/Å2 / Bits/pixel: 8
Electron beamAcceleration voltage: 100 kV / Electron source: TUNGSTEN HAIRPIN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 5.6 mm / Nominal magnification: 60000
Sample stageSpecimen holder: Standard Jeol 1200 holder / Specimen holder model: JEOL

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Image processing

DetailsThe particles were selected by manual picking
CTF correctionDetails: CTFFIND
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 20.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: EMAN, XMIPP / Number images used: 9623
Final two d classificationNumber classes: 75

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