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- EMDB-63065: Cryo-EM structure of CotVW filament, bacillus subtilis endospore ... -

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Basic information

Entry
Database: EMDB / ID: EMD-63065
TitleCryo-EM structure of CotVW filament, bacillus subtilis endospore protein
Map data
Sample
  • Complex: Helical filament of Bacillus subtilis endospore protein CotVW
    • Protein or peptide: Spore coat protein V
    • Protein or peptide: Spore coat protein W
Keywordsbacillus / spore protein / complex / filament / helical / PROTEIN FIBRIL / STRUCTURAL PROTEIN
Function / homologySpore coat protein X/V / Spore Coat Protein X and V domain / spore wall / sporulation resulting in formation of a cellular spore / Spore coat protein V / Spore coat protein W
Function and homology information
Biological speciesBacillus subtilis subsp. subtilis str. 168 (bacteria)
Methodhelical reconstruction / cryo EM / Resolution: 3.32 Å
AuthorsJo E / Kim D / Baek Y / Ha N-C
Funding support Korea, Republic Of, 1 items
OrganizationGrant numberCountry
Not funded Korea, Republic Of
CitationJournal: J Biol Chem / Year: 2025
Title: Filamentous structure of the CotVW complex, the crust proteins of the Bacillus subtilis endospore.
Authors: Eunbyul Jo / Doyeon Kim / Yeongjin Baek / Migak Park / Hyojeong Lee / Nam-Chul Ha /
Abstract: The endospores of Bacillus subtilis are encased in a multilayered protective structure comprising core, cortex, inner and outer coats, and an outermost crust. Among the proteins required for crust ...The endospores of Bacillus subtilis are encased in a multilayered protective structure comprising core, cortex, inner and outer coats, and an outermost crust. Among the proteins required for crust formation, CotV and CotW are unique to B. subtilis and are hypothesized to be instrumental in maintaining spore surface integrity. However, their structural organization and functional mechanisms remain unclear. This study determined the cryo-EM structure of the CotVW complex and revealed its filamentous helical architecture. Structural analysis showed that CotVW possesses a negatively charged surface that enables pH-dependent binding interactions. Specifically, at pH 6.0, CotVW engages in electrostatic interactions with histidine and positively charged residues, suggesting a potential regulatory mechanism influenced by the environmental pH. Our results elucidate the molecular basis of CotVW function in B. subtilis spore crust formation, highlighting its role in spore surface organization. This study advances our understanding of the spore coat architecture and may inform future research on bacterial spore resilience and structural adaptation.
History
DepositionJan 10, 2025-
Header (metadata) releaseSep 24, 2025-
Map releaseSep 24, 2025-
UpdateSep 24, 2025-
Current statusSep 24, 2025Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_63065.png

Downloads & links

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Map

FileDownload / File: emd_63065.map.gz / Format: CCP4 / Size: 59.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
0.81 Å/pix.
x 250 pix.
= 202.5 Å		0.81 Å/pix.
x 250 pix.
= 202.5 Å		0.81 Å/pix.
x 250 pix.
= 202.5 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.81 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.037
Minimum - Maximum-0.19102135 - 0.33675158
Average (Standard dev.)0.0011446197 (±0.012768693)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions250250250
Spacing250250250
CellA=B=C: 202.5 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_63065_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_63065_half_map_2.map
Projections & Slices
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Sample components

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Entire : Helical filament of Bacillus subtilis endospore protein CotVW

EntireName: Helical filament of Bacillus subtilis endospore protein CotVW
Components
  • Complex: Helical filament of Bacillus subtilis endospore protein CotVW
    • Protein or peptide: Spore coat protein V
    • Protein or peptide: Spore coat protein W

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Supramolecule #1: Helical filament of Bacillus subtilis endospore protein CotVW

SupramoleculeName: Helical filament of Bacillus subtilis endospore protein CotVW
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: The CotVW filament is composed of repeating heterodimeric units of CotV and CotW.
Source (natural)Organism: Bacillus subtilis subsp. subtilis str. 168 (bacteria)
Molecular weightTheoretical: 26.5 kDa/nm

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Macromolecule #1: Spore coat protein V

MacromoleculeName: Spore coat protein V / type: protein_or_peptide / ID: 1
Details: This sequence includes an expression tag (MGSSHHHHHHSQDP) and an additional tyrosine residue was arbitrarily added at the C-terminus for purification convenience.
Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Bacillus subtilis subsp. subtilis str. 168 (bacteria)
Molecular weightTheoretical: 16.018232 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MGSSHHHHHH SQDPMSFEEK VESLHPAIFE QLSSEFEQQI EVIDCENITI DTSHITAALS IQAFVTTMII VATQLVIADE DLADAVASE ILILDSSQIK KRTIIKIINS RNIKITLSAD EIITFVQILL QVLNSILSEL DVLY

UniProtKB: Spore coat protein V

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Macromolecule #2: Spore coat protein W

MacromoleculeName: Spore coat protein W / type: protein_or_peptide / ID: 2
Details: An additional tyrosine residue was arbitrarily added at the C-terminus for purification convenience.
Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Bacillus subtilis subsp. subtilis str. 168 (bacteria)
Molecular weightTheoretical: 12.524431 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MSDNDKFKEE LAKLPEVDPM TKMLVQNIFS KHGVTKDKMK KVSDEEKEML LNLVKDLQAK SQALIENQKK KKEEAAAQEQ KNTKPLSRR EQLIEQIRQR RKNDNNY

UniProtKB: Spore coat protein W

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statehelical array

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Sample preparation

BufferpH: 8
Component:
ConcentrationFormulaName
150.0 mMNaClsodium chloride
20.0 mMTris-HClTris(hydroxymethyl)aminomethane hydrochloride
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: TFS FALCON 4i (4k x 4k) / Number grids imaged: 1 / Number real images: 4742 / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.7 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 96000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Helical parameters - Δz: 9.52 Å
Applied symmetry - Helical parameters - Δ&Phi: 153.09 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Resolution.type: BY AUTHOR / Resolution: 3.32 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.6.0) / Number images used: 137931
CTF correctionSoftware - Name: cryoSPARC (ver. 4.6.0) / Software - details: patch ctf / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Segment selectionNumber selected: 150717 / Software - Name: cryoSPARC (ver. 4.6.0) / Software - details: auto filament picking
Startup modelType of model: NONE
Final angle assignmentType: NOT APPLICABLE / Software - Name: cryoSPARC (ver. 4.6.0)

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Atomic model buiding 1

Initial modelChain - Source name: Other / Chain - Initial model type: in silico model / Details: An initial model was generated using ModelAngelo.
RefinementSpace: REAL / Protocol: AB INITIO MODEL
Output model

PDB-9lgh:
Cryo-EM structure of CotVW filament, bacillus subtilis endospore protein

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