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- EMDB-0721: Pore structure of Iota toxin binding component (Ib) -

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Basic information

Entry
Database: EMDB / ID: EMD-0721
TitlePore structure of Iota toxin binding component (Ib)
Map data
Sample
  • Complex: Pore structure of Iota toxin binding component (Ib)
    • Protein or peptide: Iota toxin component Ib
  • Ligand: CALCIUM IONCalcium
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: 2.9 Å
AuthorsYoshida T / Yamada T
Funding support Japan, 2 items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science18K06170 Japan
Japan Society for the Promotion of Science17K15095 Japan
CitationJournal: Nat Struct Mol Biol / Year: 2020
Title: Cryo-EM structures reveal translocational unfolding in the clostridial binary iota toxin complex.
Authors: Tomohito Yamada / Toru Yoshida / Akihiro Kawamoto / Kaoru Mitsuoka / Kenji Iwasaki / Hideaki Tsuge /
Abstract: The iota toxin produced by Clostridium perfringens type E is a binary toxin comprising two independent polypeptides: Ia, an ADP-ribosyltransferase, and Ib, which is involved in cell binding and ...The iota toxin produced by Clostridium perfringens type E is a binary toxin comprising two independent polypeptides: Ia, an ADP-ribosyltransferase, and Ib, which is involved in cell binding and translocation of Ia across the cell membrane. Here we report cryo-EM structures of the translocation channel Ib-pore and its complex with Ia. The high-resolution Ib-pore structure demonstrates a similar structural framework to that of the catalytic ϕ-clamp of the anthrax protective antigen pore. However, the Ia-bound Ib-pore structure shows a unique binding mode of Ia: one Ia binds to the Ib-pore, and the Ia amino-terminal domain forms multiple weak interactions with two additional Ib-pore constriction sites. Furthermore, Ib-binding induces tilting and partial unfolding of the Ia N-terminal α-helix, permitting its extension to the ϕ-clamp gate. This new mechanism of N-terminal unfolding is crucial for protein translocation.
History
DepositionJul 30, 2019-
Header (metadata) releaseJan 15, 2020-
Map releaseJan 15, 2020-
UpdateMar 27, 2024-
Current statusMar 27, 2024Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.035
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.035
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6klx
  • Surface level: 0.035
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6klx
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_0721.png

Downloads & links

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Map

FileDownload / File: emd_0721.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.13 Å
Density
Contour LevelBy AUTHOR: 0.035 / Movie #1: 0.035
Minimum - Maximum-0.20332393 - 0.33123884
Average (Standard dev.)0.00024220705 (±0.0059767906)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 361.6 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.131.131.13
M x/y/z320320320
origin x/y/z0.0000.0000.000
length x/y/z361.600361.600361.600
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS320320320
D min/max/mean-0.2030.3310.000

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Supplemental data

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Sample components

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Entire : Pore structure of Iota toxin binding component (Ib)

EntireName: Pore structure of Iota toxin binding component (Ib)
Components
  • Complex: Pore structure of Iota toxin binding component (Ib)
    • Protein or peptide: Iota toxin component Ib
  • Ligand: CALCIUM IONCalcium

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Supramolecule #1: Pore structure of Iota toxin binding component (Ib)

SupramoleculeName: Pore structure of Iota toxin binding component (Ib) / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Clostridium perfringens (bacteria)
Molecular weightTheoretical: 520 KDa

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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: 74.37882 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: SAAWEDEDLD TDNDNIPDAY EKNGYTIKDS IAVKWNDSFA EQGYKKYVSS YLESNTAGDP YTDYQKASGS IDKAIKLEAR DPLVAAYPV VGVGMENLII STNEHASSDQ GKTVSRATTN SKTDANTVGV SISAGYQNGF TGNITTSYSH TTDNSTAVQD S NGESWNTG ...String:
SAAWEDEDLD TDNDNIPDAY EKNGYTIKDS IAVKWNDSFA EQGYKKYVSS YLESNTAGDP YTDYQKASGS IDKAIKLEAR DPLVAAYPV VGVGMENLII STNEHASSDQ GKTVSRATTN SKTDANTVGV SISAGYQNGF TGNITTSYSH TTDNSTAVQD S NGESWNTG LSINKGESAY INANVRYYNT GTAPMYKVTP TTNLVLDGET LATIKAQDNQ IGNNLSPNET YPKKGLSPLA LN TMDQFNA RLIPINYDQL KKLDSGKQIK LETTQVSGNY GTKNSQGQII TEGNSWSNYI SQIDSVSASI ILDTGSQTFE RRV AAKEQG NPEDKTPEIT IGEAIKKAFS ATKNGELLYF NGIPIDESCV ELIFDDNTSE IIKEQLKYLD DKKIYNVKLE RGMN ILIKV PSYFTNFDEY NNFPASWSNI DTKNQDGLQS VANKLSGETK IIIPMSKLKP YKRYVFSGYS KDPSTSNSIT VNIKS KEQK TDYLVPEKDY TKFSYEFETT GKDSSDIEIT LTSSGVIFLD NLSITELNST PEILKEPEIK VPSDQEILDA HNKYYA DIK LDTNTGNTYI DGIYFEPTQT NKEALDYIQK YRVEATLQYS GFKDIGTKDK EIRNYLGDQN QPKTNYINFR SYFTSGE NV MTYKKLRIYA VTPDNRELLV LSVN

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

Concentration0.38 mg/mL
BufferpH: 7.5
Component:
ConcentrationNameFormula
10.0 mMHEPES
1.0 mMcalcium chlorideCaCl2
0.01 %LMNG
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 59000
Specialist opticsSpherical aberration corrector: Microscope was modified with a Cs corrector
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
TemperatureMin: 78.9 K / Max: 78.9 K
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number grids imaged: 1 / Number real images: 2120 / Average exposure time: 84.09 sec. / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 299491
Startup modelType of model: INSILICO MODEL
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final 3D classificationSoftware - Name: RELION (ver. 3.0)
Final angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 3.0)
Final reconstructionApplied symmetry - Point group: C7 (7 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0) / Number images used: 38433
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

3j9c


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Overall B value: 46
Output model

PDB-6klx:
Pore structure of Iota toxin binding component (Ib)

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