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- EMDB-2068: Alp12 filament structure -

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Basic information

Entry
Database: EMDB / ID: EMD-2068
TitleAlp12 filament structure
Map dataReconstruction of the Alp12 filament
Sample
  • Sample: Filament structure of an actin-like protein, Alp12, in Clostridium tetani
  • Protein or peptide: Alp12
KeywordsAlp12 / actin-like protein / Clostridium tetani
Function / homologyActin-like protein, N-terminal / Actin like proteins N terminal domain / ATPase, nucleotide binding domain / Actin-like protein N-terminal domain-containing protein
Function and homology information
Biological speciesClostridium tetani (bacteria)
Methodhelical reconstruction / negative staining / Resolution: 19.7 Å
AuthorsPopp D / Narita A / Lee LJ / Ghoshdastider U / Xue B / Srinivasan R / Balasubramanian MK / Tanaka T / Robinson RC
CitationJournal: J Biol Chem / Year: 2012
Title: Novel actin-like filament structure from Clostridium tetani.
Authors: David Popp / Akihiro Narita / Lin Jie Lee / Umesh Ghoshdastider / Bo Xue / Ramanujam Srinivasan / Mohan K Balasubramanian / Toshitsugu Tanaka / Robert C Robinson /
Abstract: Eukaryotic F-actin is constructed from two protofilaments that gently wind around each other to form a helical polymer. Several bacterial actin-like proteins (Alps) are also known to form F-actin- ...Eukaryotic F-actin is constructed from two protofilaments that gently wind around each other to form a helical polymer. Several bacterial actin-like proteins (Alps) are also known to form F-actin-like helical arrangements from two protofilaments, yet with varied helical geometries. Here, we report a unique filament architecture of Alp12 from Clostridium tetani that is constructed from four protofilaments. Through fitting of an Alp12 monomer homology model into the electron microscopy data, the filament was determined to be constructed from two antiparallel strands, each composed of two parallel protofilaments. These four protofilaments form an open helical cylinder separated by a wide cleft. The molecular interactions within single protofilaments are similar to F-actin, yet interactions between protofilaments differ from those in F-actin. The filament structure and assembly and disassembly kinetics suggest Alp12 to be a dynamically unstable force-generating motor involved in segregating the pE88 plasmid, which encodes the lethal tetanus toxin, and thus a potential target for drug design. Alp12 can be repeatedly cycled between states of polymerization and dissociation, making it a novel candidate for incorporation into fuel-propelled nanobiopolymer machines.
History
DepositionApr 6, 2012-
Header (metadata) releaseApr 13, 2012-
Map releaseMay 17, 2012-
UpdateApr 20, 2016-
Current statusApr 20, 2016Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.88
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.88
  • Imaged by UCSF Chimera
  • Download
  • Surface view with fitted model
  • Atomic models: PDB-4apw
  • Surface level: 0.88
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd-2068.jpg

Downloads & links

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Map

FileDownload / File: emd_2068.map.gz / Format: CCP4 / Size: 3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationReconstruction of the Alp12 filament
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
2.68 Å/pix.
x 80 pix.
= 214.4 Å		2.68 Å/pix.
x 100 pix.
= 268. Å		2.68 Å/pix.
x 100 pix.
= 268. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX=Y=Z: 2.68 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.88 / Movie #1: 0.88
Minimum - Maximum-1.63097715 - 2.22191262
Average (Standard dev.)0.07950152 (±0.35939419)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions10010080
Spacing10010080
CellA: 268.0 Å / B: 268.0 Å / C: 214.40001 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.682.682.68
M x/y/z10010080
origin x/y/z0.0000.0000.000
length x/y/z268.000268.000214.400
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ454586
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS10010080
D min/max/mean-1.6312.2220.080

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Supplemental data

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Sample components

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Entire : Filament structure of an actin-like protein, Alp12, in Clostridiu...

EntireName: Filament structure of an actin-like protein, Alp12, in Clostridium tetani
Components
  • Sample: Filament structure of an actin-like protein, Alp12, in Clostridium tetani
  • Protein or peptide: Alp12

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Supramolecule #1000: Filament structure of an actin-like protein, Alp12, in Clostridiu...

SupramoleculeName: Filament structure of an actin-like protein, Alp12, in Clostridium tetani
type: sample / ID: 1000 / Oligomeric state: Helical filament / Number unique components: 1
Molecular weightMethod: Sequence

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Macromolecule #1: Alp12

MacromoleculeName: Alp12 / type: protein_or_peptide / ID: 1 / Oligomeric state: Filament / Recombinant expression: Yes
Source (natural)Organism: Clostridium tetani (bacteria)
Molecular weightTheoretical: 37 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria) / Recombinant plasmid: pSY5

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Experimental details

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Structure determination

Methodnegative staining
Processinghelical reconstruction
Aggregation statefilament

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Sample preparation

Concentration0.35 mg/mL
BufferpH: 7.5
Details: 300 mM KCl, 1 mM MgCl2, 0.5 mM DTT, 30 mM Hepes, 1 mM ATP
StainingType: NEGATIVE
Details: A drop of Alp12 solution was applied to grids, blotted, stained with 1 % uranyl acetate
GridDetails: 200 mesh copper grid with thin carbon support
VitrificationCryogen name: NONE / Instrument: OTHER / Method: Negatively staining

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Electron microscopy

MicroscopeHITACHI H7600
DateSep 6, 2011
Image recordingCategory: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: ZEISS SCAI / Digitization - Sampling interval: 7 µm / Number real images: 31 / Bits/pixel: 12
Electron beamAcceleration voltage: 100 kV / Electron source: LAB6
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 5.0 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 40000
Sample stageSpecimen holder model: SIDE ENTRY, EUCENTRIC

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Image processing

DetailsThe particles were aligned using Eos.
Final reconstructionApplied symmetry - Helical parameters - Δz: 42.88 Å
Applied symmetry - Helical parameters - Δ&Phi: 13.24 °
Applied symmetry - Helical parameters - Axial symmetry: D1 (2x1 fold dihedral)
Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 19.7 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: EOS
CTF correctionDetails: Each scanned image

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Atomic model buiding 1

Initial modelPDB ID:

3js6

DetailsProtocol: X-RAY
RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-4apw:
Alp12 filament structure

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