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- EMDB-1149: Dodecameric structure of the small heat shock protein Acr1 from M... -

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Basic information

Entry
Database: EMDB / ID: EMD-1149
TitleDodecameric structure of the small heat shock protein Acr1 from Mycobacterium tuberculosis.
Map dataNegative stain EM map of dodecamer of M.tuberculosis Acr1
Sample
  • Sample: Recombinant protein Acr1 From M.Tuberculosis made in E.coli.
  • Protein or peptide: Acr1
Function / homology
Function and homology information


response to salt stress / response to hydrogen peroxide / protein homooligomerization / : / unfolded protein binding / protein folding / protein complex oligomerization / response to heat / cytoplasm
Similarity search - Function
: / Hsp20/alpha crystallin family / Small heat shock protein (sHSP) domain profile. / Alpha crystallin/Hsp20 domain / Alpha crystallin/Hsp20 domain / HSP20-like chaperone
Similarity search - Domain/homology
16.9 kDa class I heat shock protein 2
Similarity search - Component
Biological speciesMycobacterium tuberculosis (bacteria)
Methodsingle particle reconstruction / negative staining / Resolution: 16.5 Å
AuthorsKennaway CK / Benesch JL / Gohlke U / Wang L / Robinson CV / Orlova EV / Saibil HR / Saibi HR / Keep NH
CitationJournal: J Biol Chem / Year: 2005
Title: Dodecameric structure of the small heat shock protein Acr1 from Mycobacterium tuberculosis.
Authors: Christopher K Kennaway / Justin L P Benesch / Ulrich Gohlke / Luchun Wang / Carol V Robinson / Elena V Orlova / Helen R Saibil / Nicholas H Keep /
Abstract: Small heat shock proteins are a ubiquitous and diverse family of stress proteins that have in common an alpha-crystallin domain. Mycobacterium tuberculosis has two small heat shock proteins, Acr1 ...Small heat shock proteins are a ubiquitous and diverse family of stress proteins that have in common an alpha-crystallin domain. Mycobacterium tuberculosis has two small heat shock proteins, Acr1 (alpha-crystallin-related protein 1, or Hsp16.3/16-kDa antigen) and Acr2 (HrpA), both of which are highly expressed under different stress conditions. Small heat shock proteins form large oligomeric assemblies and are commonly polydisperse. Nanoelectrospray mass spectrometry showed that Acr2 formed a range of oligomers composed of dimers and tetramers, whereas Acr1 was a dodecamer. Electron microscopy of Acr2 showed a variety of particle sizes. Using three-dimensional analysis of negative stain electron microscope images, we have shown that Acr1 forms a tetrahedral assembly with 12 polypeptide chains. The atomic structure of a related alpha-crystallin domain dimer was docked into the density to build a molecular structure of the dodecameric Acr1 complex. Along with the differential regulation of these two proteins, the differences in their quaternary structures demonstrated here supports their distinct functional roles.
History
DepositionAug 5, 2005-
Header (metadata) releaseAug 10, 2005-
Map releaseFeb 9, 2006-
UpdateMay 26, 2011-
Current statusMay 26, 2011Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.25
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.25
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-2byu
  • Surface level: 0.25
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

1149.gif

Downloads & links

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Map

FileDownload / File: emd_1149.map.gz / Format: CCP4 / Size: 1001 KB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationNegative stain EM map of dodecamer of M.tuberculosis Acr1
Voxel sizeX=Y=Z: 3.3333 Å
Density
Contour Level1: 0.0116 / Movie #1: 0.25
Minimum - Maximum-0.278048 - 1.02549
Average (Standard dev.)0.011189 (±0.0851497)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderYXZ
Origin-31-32-32
Dimensions646464
Spacing646464
CellA=B=C: 213.331 Å
α=β=γ: 90 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z3.3332968753.3332968753.333296875
M x/y/z646464
origin x/y/z0.0000.0000.000
length x/y/z213.331213.331213.331
α/β/γ90.00090.00090.000
start NX/NY/NZ-31-32-32
NX/NY/NZ646464
MAP C/R/S213
start NC/NR/NS-32-31-32
NC/NR/NS646464
D min/max/mean-0.2781.0250.011

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Supplemental data

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Sample components

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Entire : Recombinant protein Acr1 From M.Tuberculosis made in E.coli.

EntireName: Recombinant protein Acr1 From M.Tuberculosis made in E.coli.
Components
  • Sample: Recombinant protein Acr1 From M.Tuberculosis made in E.coli.
  • Protein or peptide: Acr1

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Supramolecule #1000: Recombinant protein Acr1 From M.Tuberculosis made in E.coli.

SupramoleculeName: Recombinant protein Acr1 From M.Tuberculosis made in E.coli.
type: sample / ID: 1000
Details: Sample was monodisperse by mass spectrometry and light scattering
Oligomeric state: dodecamer tetrahedral symmetry / Number unique components: 1
Molecular weightExperimental: 196 KDa / Theoretical: 196 KDa / Method: Mass spectrometry

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Macromolecule #1: Acr1

MacromoleculeName: Acr1 / type: protein_or_peptide / ID: 1 / Name.synonym: Hsp16.3 / Details: monomer / Number of copies: 12 / Oligomeric state: dodecamer / Recombinant expression: Yes
Source (natural)Organism: Mycobacterium tuberculosis (bacteria) / Strain: H37Rv / synonym: TB
Molecular weightExperimental: 18.5 KDa / Theoretical: 18.5 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli) / Recombinant plasmid: pET15b
SequenceInterPro: Alpha crystallin/Hsp20 domain

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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

Concentration0.01 mg/mL
BufferpH: 7 / Details: 50 mM Tris pH 7
StainingType: NEGATIVE / Details: Two 5 ul aliquots of 2% uranyl acetate.
GridDetails: 400 mesh copper grid with carbon film
VitrificationCryogen name: NONE / Chamber temperature: 293 K

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Electron microscopy

MicroscopeFEI TECNAI 12
Electron beamAcceleration voltage: 120 kV / Electron source: TUNGSTEN HAIRPIN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.0 mm / Nominal defocus max: 0.6 µm / Nominal defocus min: 0.39 µm / Nominal magnification: 42000
Sample stageSpecimen holder: side entry / Specimen holder model: OTHER
TemperatureAverage: 293 K
Alignment procedureLegacy - Astigmatism: corrected at 100k magnification
DateMar 26, 2004
Image recordingCategory: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: ZEISS SCAI / Digitization - Sampling interval: 14 µm / Number real images: 18 / Average electron dose: 10 e/Å2 / Od range: 1 / Bits/pixel: 8

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Image processing

Final two d classificationNumber classes: 400
Final reconstructionApplied symmetry - Point group: T (tetrahedral) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 16.5 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: IMAGIC
Details: Exact filtered back projection. Map made from 400 out of 600 class averages.
Number images used: 6123

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Atomic model buiding 1

Initial modelPDB ID:

1gme


Chain - #0 - Chain ID: A / Chain - #1 - Chain ID: B
SoftwareName: URO
DetailsPDBEntryID_givenInChain. Protocol: rigid body. One dimer of 1gme A and B, residues 43-137 and 146-151, was fitted in URO and tetrahedral symmetry was used to generate the other five dimers.
RefinementSpace: RECIPROCAL / Protocol: RIGID BODY FIT / Target criteria: Density correlation
Output model

PDB-2byu:
Negative stain EM reconstruction of M.tuberculosis Acr1(Hsp 16.3) fitted with wheat sHSP dimer

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