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- EMDB-39544: Iota toxin Ib pore serine-clamp mutant -

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Basic information

Entry
Database: EMDB / ID: EMD-39544
TitleIota toxin Ib pore serine-clamp mutant
Map data
Sample
  • Complex: Iota toxin Ib pore serine-clamp mutant
    • Protein or peptide: Iota toxin component Ib
  • Ligand: CALCIUM ION
KeywordsBacterial binary toxin / Protein translocation channel / ADP-ribosylation / Toxin
Function / homology
Function and homology information


protein homooligomerization / extracellular region
Similarity search - Function
Bacterial exotoxin B / Protective antigen, heptamerisation domain / Protective antigen, Ca-binding domain / Clostridial binary toxin B/anthrax toxin PA, domain 3 / Protective antigen, heptamerisation domain superfamily / Clostridial binary toxin B/anthrax toxin PA Ca-binding domain / Clostridial binary toxin B/anthrax toxin PA domain 2 / Clostridial binary toxin B/anthrax toxin PA domain 3 / PA14 / PA14 domain
Similarity search - Domain/homology
Iota toxin component Ib
Similarity search - Component
Biological speciesClostridium perfringens (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.36 Å
AuthorsNinomiya Y / Yoshida T / Yamada T / Kishikawa J / Tsuge H
Funding support Japan, 1 items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS)21H02452 Japan
CitationJournal: Commun Biol / Year: 2025
Title: Serine clamp of Clostridium perfringens binary toxin BECb (CPILEb)-pore confers cytotoxicity and enterotoxicity.
Authors: Toru Yoshida / Chie Monma / Yuki Ninomiya / Sotaro Takiguchi / Shoko Fujita / Yuto Uchida / Noriaki Sakoda / Vladimir A Karginov / Jun-Ichi Kishikawa / Tomohito Yamada / Ryuji Kawano / Hideaki Tsuge /
Abstract: BEC (CPILE) is a virulence factor of the pathogen, Clostridium perfringens, which has caused foodborne outbreaks in Japan. BEC is a binary toxin that comprises the enzymatic A-component (BECa) and ...BEC (CPILE) is a virulence factor of the pathogen, Clostridium perfringens, which has caused foodborne outbreaks in Japan. BEC is a binary toxin that comprises the enzymatic A-component (BECa) and the B-component (BECb); the latter forms a membrane pore to translocate the A-component into target cells. Although BEC differs from other binary toxins in that the B-component alone shows enterotoxic activity, the reason for this remains unclear. We focus on the narrowest region of BECb-pore formed by not phenylalanine residues conserved in other binary toxins including iota toxin B-component (Ib) but serine residues. Comparisons between BECb and BECb (S405F) where the serine residue forming the narrowest region is substituted to the phenylalanine residue reveal that the serine residue is responsible for both cytotoxicity and enterotoxic activity. Though attempts to prepare the BECb-pore were unsuccessful, we reveal the cryo-EM structure of Ib (F454S) where the phenylalanine residue forming the narrowest region is substituted to the serine residue as a surrogate of BECb. Furthermore, Ib (F454S) increases current conductance to nine times that of Ib due to the larger pore diameter and the hydrophilic nature. These results suggest that BECb functions as a pore-forming toxin and as a translocation channel for BECa.
History
DepositionMar 21, 2024-
Header (metadata) releaseMar 26, 2025-
Map releaseMar 26, 2025-
UpdateAug 13, 2025-
Current statusAug 13, 2025Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_39544.png

Downloads & links

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Map

FileDownload / File: emd_39544.map.gz / Format: CCP4 / Size: 347.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.68 Å/pix.
x 450 pix.
= 303.75 Å		0.68 Å/pix.
x 450 pix.
= 303.75 Å		0.68 Å/pix.
x 450 pix.
= 303.75 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.675 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.16
Minimum - Maximum-0.5295337 - 1.3181643
Average (Standard dev.)0.0010923586 (±0.037887815)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions450450450
Spacing450450450
CellA=B=C: 303.75 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_39544_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_39544_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram

Histogram (log scale)

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Sample components

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Entire : Iota toxin Ib pore serine-clamp mutant

EntireName: Iota toxin Ib pore serine-clamp mutant
Components
  • Complex: Iota toxin Ib pore serine-clamp mutant
    • Protein or peptide: Iota toxin component Ib
  • Ligand: CALCIUM ION

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Supramolecule #1: Iota toxin Ib pore serine-clamp mutant

SupramoleculeName: Iota toxin Ib pore serine-clamp mutant / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Clostridium perfringens (bacteria)

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Macromolecule #1: Iota toxin component Ib

MacromoleculeName: Iota toxin component Ib / type: protein_or_peptide / ID: 1 / Number of copies: 7 / Enantiomer: LEVO
Source (natural)Organism: Clostridium perfringens (bacteria)
Molecular weightTheoretical: 58.241887 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: EDLDTDNDNI PDAYEKNGYT IKDSIAVKWN DSFAEQGYKK YVSSYLESNT AGDPYTDYQK ASGSIDKAIK LEARDPLVAA YPVVGVGME NLIISTNEHA SSDQGKTVSR ATTNSKTDAN TVGVSISAGY QNGFTGNITT SYSHTTDNST AVQDSNGESW N TGLSINKG ...String:
EDLDTDNDNI PDAYEKNGYT IKDSIAVKWN DSFAEQGYKK YVSSYLESNT AGDPYTDYQK ASGSIDKAIK LEARDPLVAA YPVVGVGME NLIISTNEHA SSDQGKTVSR ATTNSKTDAN TVGVSISAGY QNGFTGNITT SYSHTTDNST AVQDSNGESW N TGLSINKG ESAYINANVR YYNTGTAPMY KVTPTTNLVL DGETLATIKA QDNQIGNNLS PNETYPKKGL SPLALNTMDQ SN ARLIPIN YDQLKKLDSG KQIKLETTQV SGNYGTKNSQ GQIITEGNSW SNYISQIDSV SASIILDTGS QTFERRVAAK EQG NPEDKT PEITIGEAIK KAFSATKNGE LLYFNGIPID ESCVELIFDD NTSEIIKEQL KYLDDKKIYN VKLERGMNIL IKVP SYFTN FDEYNNFPAS WSNIDTKNQD GLQSVANKLS GETKIIIPMS KLKPYKRYVF SGYSKDPSTS NSITVNIKSK EQKTD YLVP EKDYTKFSYE FETTGKDSSD IEITLTSSGV IFLDNLSITE LNST

UniProtKB: Iota toxin component Ib

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Macromolecule #2: CALCIUM ION

MacromoleculeName: CALCIUM ION / type: ligand / ID: 2 / Number of copies: 14 / Formula: CA
Molecular weightTheoretical: 40.078 Da

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.669 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
10.0 mMC8H18N2O4S2-[4-(2-Hydroxyethyl)-1-piperazinyl]ethanesulfonic acid
1.0 mMCaCl2calcium chloride
0.003 % (w/v)C47H88O22Lauryl Maltose Neopentyl Glycol

Details: 10mM HEPES pH 7.5, 1mM CaCl2, 0.003% (w/v) LMNG
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number real images: 7080 / Average exposure time: 2.89 sec. / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 0.0938 mm / Nominal defocus max: 1.8 µm / Nominal defocus min: 0.6 µm
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1379432
CTF correctionSoftware - Name: cryoSPARC (ver. 3.3.2) / Software - details: Patch CTF estimation (multi) / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: OTHER
Final reconstructionApplied symmetry - Point group: C7 (7 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.36 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.3.2) / Software - details: Homogenious refinement / Number images used: 190663
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3.2)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3.2)
Final 3D classificationNumber classes: 150 / Software - Name: cryoSPARC (ver. 3.3.2)
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-8yrm:
Iota toxin Ib pore serine-clamp mutant

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