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- EMDB-7943: Mtb ClpB in complex with ATPgammaS and casein, Conformer 2 -

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Basic information

Entry
Database: EMDB / ID: 7943
TitleMtb ClpB in complex with ATPgammaS and casein, Conformer 2
Map dataMtb ClpB in complex with ATPgammaS and casein, Conformer 2
SampleMtb ClpB in complex with ATPgammaS and casein:
Chaperone protein ClpB / casein polyAlanine model / (ligand) x 2
Function / homologyClp ATPase, C-terminal / ClpA/B, conserved site 1 / AAA domain (Cdc48 subfamily) / Clp amino terminal domain, pathogenicity island component / ATPase family associated with various cellular activities (AAA) / Clp, N-terminal domain superfamily / ClpA/B, conserved site 2 / P-loop containing nucleoside triphosphate hydrolase / Chaperonins clpA/B signature 2. / Chaperonin ClpB ...Clp ATPase, C-terminal / ClpA/B, conserved site 1 / AAA domain (Cdc48 subfamily) / Clp amino terminal domain, pathogenicity island component / ATPase family associated with various cellular activities (AAA) / Clp, N-terminal domain superfamily / ClpA/B, conserved site 2 / P-loop containing nucleoside triphosphate hydrolase / Chaperonins clpA/B signature 2. / Chaperonin ClpB / Chaperonins clpA/B signature 1. / Clp, N-terminal / ATPase, AAA-type, core / AAA+ ATPase domain / ClpA/B family / C-terminal, D2-small domain, of ClpB protein / protein metabolic process / response to heat / protein refolding / ATP binding / cytoplasm / Chaperone protein ClpB
Function and homology information
SourceMycobacterium tuberculosis (bacteria) / Bos taurus (cattle)
Methodsingle particle reconstruction / cryo EM / 3.9 Å resolution
AuthorsYu HJ / Li HL
CitationJournal: Proc. Natl. Acad. Sci. U.S.A. / Year: 2018
Title: ATP hydrolysis-coupled peptide translocation mechanism of ClpB.
Authors: Hongjun Yu / Tania J Lupoli / Amanda Kovach / Xing Meng / Gongpu Zhao / Carl F Nathan / Huilin Li
Abstract: The protein disaggregase ClpB hexamer is conserved across evolution and has two AAA+-type nucleotide-binding domains, NBD1 and NBD2, in each protomer. In (), ClpB facilitates asymmetric distribution ...The protein disaggregase ClpB hexamer is conserved across evolution and has two AAA+-type nucleotide-binding domains, NBD1 and NBD2, in each protomer. In (), ClpB facilitates asymmetric distribution of protein aggregates during cell division to help the pathogen survive and persist within the host, but a mechanistic understanding has been lacking. Here we report cryo-EM structures at 3.8- to 3.9-Å resolution of ClpB bound to a model substrate, casein, in the presence of the weakly hydrolyzable ATP mimic adenosine 5'-[γ-thio]triphosphate. ClpB existed in solution in two closed-ring conformations, conformers 1 and 2. In both conformers, the 12 pore-loops on the 12 NTDs of the six protomers (P1-P6) were arranged similarly to a staircase around the bound peptide. Conformer 1 is a low-affinity state in which three of the 12 pore-loops (the protomer P1 NBD1 and NBD2 loops and the protomer P2 NBD1 loop) are not engaged with peptide. Conformer 2 is a high-affinity state because only one pore-loop (the protomer P2 NBD1 loop) is not engaged with the peptide. The resolution of the two conformations, along with their bound substrate peptides and nucleotides, enabled us to propose a nucleotide-driven peptide translocation mechanism of a bacterial ClpB that is largely consistent with several recent unfoldase structures, in particular with the eukaryotic Hsp104. However, whereas Hsp104's two NBDs move in opposing directions during one step of peptide translocation, in Mtb ClpB the two NBDs move only in the direction of translocation.
Validation ReportPDB-ID: 6djv

SummaryFull reportAbout validation report
DateDeposition: May 26, 2018 / Header (metadata) release: Jul 11, 2018 / Map release: Sep 26, 2018 / Last update: Oct 24, 2018

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.04
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.04
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-6djv
  • Surface level: 0.04
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

Fileemd_7943.map.gz (map file in CCP4 format, 99589 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
292 pix
1.07 Å/pix.
= 313.608 Å
292 pix
1.07 Å/pix.
= 313.608 Å
292 pix
1.07 Å/pix.
= 313.608 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.074 Å
Density
Contour Level:0.033 (by author), 0.04 (movie #1):
Minimum - Maximum-0.08667084 - 0.21426883
Average (Standard dev.)0.00034849244 (0.0068144323)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions292292292
Origin0.00.00.0
Limit291.0291.0291.0
Spacing292292292
CellA=B=C: 313.608 Å
α=β=γ: 90.0 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0741.0741.074
M x/y/z292292292
origin x/y/z0.0000.0000.000
length x/y/z313.608313.608313.608
α/β/γ90.00090.00090.000
start NX/NY/NZ
NX/NY/NZ
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS292292292
D min/max/mean-0.0870.2140.000

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Supplemental data

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Sample components

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Entire Mtb ClpB in complex with ATPgammaS and casein

EntireName: Mtb ClpB in complex with ATPgammaS and casein / Number of components: 5

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Component #1: protein, Mtb ClpB in complex with ATPgammaS and casein

ProteinName: Mtb ClpB in complex with ATPgammaS and casein / Recombinant expression: No
SourceSpecies: Mycobacterium tuberculosis (bacteria)

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Component #2: protein, Chaperone protein ClpB

ProteinName: Chaperone protein ClpB / Number of Copies: 6 / Recombinant expression: No
MassTheoretical: 92.688281 kDa
SourceSpecies: Mycobacterium tuberculosis (bacteria)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #3: protein, casein polyAlanine model

ProteinName: casein polyAlanine model / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 1.866038 kDa
SourceSpecies: Bos taurus (cattle)

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Component #4: ligand, PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER

LigandName: PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER / Number of Copies: 10 / Recombinant expression: No
MassTheoretical: 0.523247 kDa

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Component #5: ligand, ADENOSINE-5'-DIPHOSPHATE

LigandName: ADENOSINE-5'-DIPHOSPHATE / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 0.427201 kDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: particle / Method: cryo EM
Sample solutionpH: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 1.7 e/Å2 / Illumination mode: FLOOD BEAM
LensImaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C1 (asymmetric) / Number of projections: 76666
3D reconstructionSoftware: RELION / Resolution: 3.9 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot
(resolution estimation)

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Atomic model buiding

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