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- PDB-9cmh: Cryo-EM structure of human claudin-4 complex with Clostridium per... -

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Basic information

Entry
Database: PDB / ID: 9cmh
TitleCryo-EM structure of human claudin-4 complex with Clostridium perfringens enterotoxin
Components
  • Claudin-4
  • Heat-labile enterotoxin B chain
KeywordsMEMBRANE PROTEIN / claudin / enterotoxin
Function / homology
Function and homology information


paracellular transport / calcium-independent cell-cell adhesion via plasma membrane cell-adhesion molecules / Tight junction interactions / bicellular tight junction assembly / apicolateral plasma membrane / regulation of cell morphogenesis / tight junction / positive regulation of wound healing / renal absorption / chloride channel activity ...paracellular transport / calcium-independent cell-cell adhesion via plasma membrane cell-adhesion molecules / Tight junction interactions / bicellular tight junction assembly / apicolateral plasma membrane / regulation of cell morphogenesis / tight junction / positive regulation of wound healing / renal absorption / chloride channel activity / chloride channel complex / bicellular tight junction / lateral plasma membrane / establishment of skin barrier / basal plasma membrane / response to progesterone / female pregnancy / circadian rhythm / transmembrane signaling receptor activity / cell-cell junction / toxin activity / cell adhesion / positive regulation of cell migration / apical plasma membrane / structural molecule activity / extracellular region / identical protein binding / plasma membrane
Similarity search - Function
Claudin-4 / Claudin / Claudin, conserved site / Claudin family signature. / Clostridium enterotoxin / Clostridium enterotoxin / PMP-22/EMP/MP20/Claudin family / PMP-22/EMP/MP20/Claudin superfamily
Similarity search - Domain/homology
Claudin-4 / Heat-labile enterotoxin B chain
Similarity search - Component
Biological speciesHomo sapiens (human)
Clostridium perfringens (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4 Å
AuthorsVecchio, A.J.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM138368 United States
CitationJournal: Structure / Year: 2024
Title: Cryo-EM structures of Clostridium perfringens enterotoxin bound to its human receptor, claudin-4.
Authors: Sewwandi S Rathnayake / Satchal K Erramilli / Anthony A Kossiakoff / Alex J Vecchio /
Abstract: Clostridium perfringens enterotoxin (CpE) causes prevalent and deadly gastrointestinal disorders. CpE binds to receptors called claudins on the apical surfaces of small intestinal epithelium. ...Clostridium perfringens enterotoxin (CpE) causes prevalent and deadly gastrointestinal disorders. CpE binds to receptors called claudins on the apical surfaces of small intestinal epithelium. Claudins normally regulate paracellular transport but are hijacked from doing so by CpE and are instead led to form claudin/CpE complexes. Claudin/CpE complexes are the building blocks of oligomeric β-barrel pores that penetrate the plasma membrane and induce gut cytotoxicity. Here, we present the structures of CpE in complex with its native claudin receptor in humans, claudin-4, using cryogenic electron microscopy. The structures reveal the architecture of the claudin/CpE complex, the residues used in binding, the orientation of CpE relative to the membrane, and CpE-induced changes to claudin-4. Further, structures and modeling allude to the biophysical procession from claudin/CpE complexes to cytotoxic β-barrel pores during pathogenesis. In full, this work proposes a model of claudin/CpE assembly and provides strategies to obstruct its formation to treat CpE diseases.
History
DepositionJul 15, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 24, 2024Provider: repository / Type: Initial release
Revision 1.1Oct 23, 2024Group: Data collection / Structure summary
Category: em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _em_admin.last_update
Revision 1.2Oct 30, 2024Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID / _citation_author.name / _em_admin.last_update
Revision 1.3Nov 27, 2024Group: Data collection / Database references / Category: citation / em_admin
Item: _citation.journal_volume / _citation.page_first / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Claudin-4
B: Heat-labile enterotoxin B chain


Theoretical massNumber of molelcules
Total (without water)55,2352
Polymers55,2352
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable, gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Claudin-4 / Clostridium perfringens enterotoxin receptor / CPE-R / CPE-receptor / Williams-Beuren syndrome ...Clostridium perfringens enterotoxin receptor / CPE-R / CPE-receptor / Williams-Beuren syndrome chromosomal region 8 protein


Mass: 22234.332 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CLDN4, CPER, CPETR1, WBSCR8 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: O14493
#2: Protein Heat-labile enterotoxin B chain


Mass: 33000.582 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Clostridium perfringens (bacteria) / Gene: cpe / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P01558
Has protein modificationY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Human claudin-4 complex with Clostridium perfringens enterotoxin
Type: COMPLEX / Details: 2-protein complex / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 0.055 MDa / Experimental value: NO
Source (natural)Organism: other entries (others)
Source (recombinant)Organism: Trichoplusia ni (cabbage looper)
Buffer solutionpH: 7.4 / Details: 20 mM Hepes pH 7.4, 100 mM NaCl, and 0.003% LMNG
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMHepes pH 7.41
2100 mMNaClNaCl1
30.003 %LMNG1
SpecimenConc.: 8 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: glow-discharged for 60 seconds at 15 mA using a Pelco easiGlow (Ted Pella Inc) instrument.
Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3
VitrificationInstrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 129000 X / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm / Alignment procedure: BASIC
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 60 e/Å2 / Film or detector model: GATAN K3 BIOCONTINUUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 7100
EM imaging opticsEnergyfilter name: GIF Bioquantum

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Processing

EM software
IDNameVersionCategory
1cryoSPARC4.4particle selection
2SerialEMimage acquisition
4cryoSPARC4.4CTF correction
7UCSF Chimeramodel fitting
9cryoSPARC4.4initial Euler assignment
10cryoSPARC4.4final Euler assignment
11cryoSPARC4.4classification
12cryoSPARC4.43D reconstruction
13PHENIX1.20.1_4487:model refinement
CTF correctionType: NONE
Particle selectionNum. of particles selected: 2504188
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 44683 / Algorithm: FOURIER SPACE / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL
Atomic model building
IDPDB-ID 3D fitting-IDAccession codeInitial refinement model-IDSource nameType
17kp4

7kp4

17kp41PDBexperimental model
28u5f

8u5f

18u5f2PDBexperimental model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0043619
ELECTRON MICROSCOPYf_angle_d0.7984923
ELECTRON MICROSCOPYf_dihedral_angle_d4.898495
ELECTRON MICROSCOPYf_chiral_restr0.046579
ELECTRON MICROSCOPYf_plane_restr0.004618

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