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- EMDB-4062: MamK double helical filament at 3.6 A resolution -

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Basic information

Entry
Database: EMDB / ID: EMD-4062
TitleMamK double helical filament at 3.6 A resolution
Map dataNone
Sample
  • Complex: MamK
    • Protein or peptide: Actin-like ATPase
  • Ligand: MAGNESIUM ION
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
Function / homology
Function and homology information


magnetosome assembly / magnetosome membrane / Hydrolases; Acting on acid anhydrides; In phosphorus-containing anhydrides / cytoskeleton / GTP binding / ATP hydrolysis activity / ATP binding / metal ion binding / cytoplasm
Similarity search - Function
MreB/Mbl protein / Actin family / Actin / ATPase, nucleotide binding domain
Similarity search - Domain/homology
Actin-like protein MamK
Similarity search - Component
Biological speciesMagnetospirillum magneticum AMB-1 (bacteria)
Methodhelical reconstruction / cryo EM / Resolution: 3.64 Å
AuthorsLowe J
Funding support United Kingdom, 2 items
OrganizationGrant numberCountry
Medical Research Council (United Kingdom)U105184326 United Kingdom
Wellcome Trust095514/Z/11/Z United Kingdom
CitationJournal: Proc Natl Acad Sci U S A / Year: 2016
Title: X-ray and cryo-EM structures of monomeric and filamentous actin-like protein MamK reveal changes associated with polymerization.
Authors: Jan Löwe / Shaoda He / Sjors H W Scheres / Christos G Savva /
Abstract: Magnetotactic bacteria produce iron-rich magnetic nanoparticles that are enclosed by membrane invaginations to form magnetosomes so they are able to sense and act upon Earth's magnetic field. In ...Magnetotactic bacteria produce iron-rich magnetic nanoparticles that are enclosed by membrane invaginations to form magnetosomes so they are able to sense and act upon Earth's magnetic field. In Magnetospirillum and other magnetotactic bacteria, to combine their magnetic moments, magnetosomes align along filaments formed by a bacterial actin homolog, MamK. Here, we present the crystal structure of a nonpolymerizing mutant of MamK from Magnetospirillum magneticum AMB-1 at 1.8-Å resolution, revealing its close similarity to actin and MreB. The crystals contain AMPPNP-bound monomeric MamK in two different conformations. To investigate conformational changes associated with polymerization, we used unmodified MamK protein and cryo-EM with helical 3D reconstruction in RELION to obtain a density map and a fully refined atomic model of MamK in filamentous form at 3.6-Å resolution. The filament is parallel (polar) double-helical, with a rise of 52.2 Å and a twist of 23.8°. As shown previously and unusually for actin-like filaments, the MamK subunits from each of the two strands are juxtaposed, creating an additional twofold axis along the filament. Compared with monomeric MamK, ADP-bound MamK in the filament undergoes a conformational change, rotating domains I and II against each other to further close the interdomain cleft between subdomains IB and IIB. The domain movement causes several loops to close around the nucleotide-binding pocket. Glu-143, a key residue for catalysis coordinating the magnesium ion, moves closer, presumably switching nucleotide hydrolysis upon polymerization-one of the hallmarks of cytomotive filaments of the actin type.
History
DepositionJul 21, 2016-
Header (metadata) releaseOct 12, 2016-
Map releaseNov 16, 2016-
UpdateOct 23, 2019-
Current statusOct 23, 2019Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.155
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.155
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-5ljv
  • Surface level: 0.155
  • Imaged by UCSF Chimera
  • Download
  • Surface view with fitted model
  • Atomic models: PDB-5ljv
  • Surface level: 0.155
  • Imaged by UCSF Chimera
  • Download
  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-5ljv
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_4062.map.gz / Format: CCP4 / Size: 83.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationNone
Voxel sizeX=Y=Z: 1.34 Å
Density
Contour LevelBy AUTHOR: 0.155 / Movie #1: 0.155
Minimum - Maximum-0.418076 - 0.82236314
Average (Standard dev.)-0.0001257372 (±0.02321098)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions280280280
Spacing280280280
CellA=B=C: 375.2 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.341.341.34
M x/y/z280280280
origin x/y/z0.0000.0000.000
length x/y/z375.200375.200375.200
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS280280280
D min/max/mean-0.4180.822-0.000

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Supplemental data

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Sample components

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Entire : MamK

EntireName: MamK
Components
  • Complex: MamK
    • Protein or peptide: Actin-like ATPase
  • Ligand: MAGNESIUM ION
  • Ligand: ADENOSINE-5'-DIPHOSPHATE

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Supramolecule #1: MamK

SupramoleculeName: MamK / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Magnetospirillum magneticum AMB-1 (bacteria)
Recombinant expressionOrganism: Escherichia coli (E. coli) / Recombinant plasmid: pHis17

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Macromolecule #1: Actin-like ATPase

MacromoleculeName: Actin-like ATPase / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Magnetospirillum magneticum AMB-1 (bacteria)
Molecular weightTheoretical: 37.642152 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MSEGEGQAKN RLFLGVDLGT SHTAVMSSRG KKFLLKSVVG YPKDVIGLKL LGRPYVVGDE AFEMRSYLDI RYPLQDGVLS EISDRDIEV ARHLLTHVVK SAEPGPNDEI CAVIGVPARA SAANKALLLK MAQEVVHTAL VVSEPFMVGY GLDKLINTII V DIGAGTTD ...String:
MSEGEGQAKN RLFLGVDLGT SHTAVMSSRG KKFLLKSVVG YPKDVIGLKL LGRPYVVGDE AFEMRSYLDI RYPLQDGVLS EISDRDIEV ARHLLTHVVK SAEPGPNDEI CAVIGVPARA SAANKALLLK MAQEVVHTAL VVSEPFMVGY GLDKLINTII V DIGAGTTD ICALKGTVPG PEDQVTLTKA GNYVDERLQN AILERHPELQ MNVNVACAVK EQFSFVGTPT EVASFEFRAA GK PVRADVT EPVKIACEAL MPDIIESIET LLRSFQPEYQ ATVLQNIVFA GGGSRIRGLA AYVKEKLRPF GDANVTCVKD PTF DGCRGA LRLAEELPPQ YWRQLGDVSG S

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Macromolecule #2: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 2 / Number of copies: 6 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #3: ADENOSINE-5'-DIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-DIPHOSPHATE / type: ligand / ID: 3 / Number of copies: 6 / Formula: ADP
Molecular weightTheoretical: 427.201 Da
Chemical component information

ChemComp-ADP:
ADENOSINE-5'-DIPHOSPHATE / ADP, energy-carrying molecule*YM / Adenosine diphosphate

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statefilament

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Sample preparation

Concentration1.5 mg/mL
BufferpH: 7.4
GridModel: Quantifoil / Material: COPPER/RHODIUM / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 283 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI POLARA 300
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Calibrated defocus max: 3.0 µm / Calibrated defocus min: 1.1 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Image recordingFilm or detector model: FEI FALCON II (4k x 4k) / Detector mode: INTEGRATING / Digitization - Dimensions - Width: 4000 pixel / Digitization - Dimensions - Height: 4000 pixel / Number grids imaged: 3 / Number real images: 1665 / Average exposure time: 1.5 sec. / Average electron dose: 40.0 e/Å2
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: Gctf
Final angle assignmentType: NOT APPLICABLE / Software - Name: RELION
Final reconstructionApplied symmetry - Helical parameters - Δz: 52.15 Å
Applied symmetry - Helical parameters - Δ&Phi: 23.77 °
Applied symmetry - Helical parameters - Axial symmetry: C2 (2 fold cyclic)
Resolution.type: BY AUTHOR / Resolution: 3.64 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 596427
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: RECIPROCAL / Protocol: OTHER / Target criteria: R-factor
Output model

PDB-5ljv:
MamK double helical filament

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