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- PDB-7p3r: Helical structure of the toxin MakA from Vibrio cholera -

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Basic information

Entry
Database: PDB / ID: 7p3r
TitleHelical structure of the toxin MakA from Vibrio cholera
ComponentsMakA tetramer
KeywordsTOXIN / Pore-forming toxin / Vibrio cholerae
Function / homologyHemolysin BL-binding component / Bacillus haemolytic enterotoxin (HBL) / : / Hemolysin E; Chain: A; / Hemolysin E; Chain: A; - #10 / Up-down Bundle / Mainly Alpha / membrane / Non-hemolytic enterotoxin lytic component L1
Function and homology information
Biological speciesVibrio cholerae O1 biovar El Tor str. N16961 (bacteria)
MethodELECTRON MICROSCOPY / helical reconstruction / cryo EM / Resolution: 3.65 Å
AuthorsBerg, A. / Nadeem, A. / Uhlin, B.E. / Wai, S.N. / Barandun, J.
Funding support Sweden, European Union, 4items
OrganizationGrant numberCountry
Swedish Research Council2019-02011 Sweden
European Research Council (ERC)948655European Union
Swedish Research Council2018-02914 Sweden
Swedish Research Council2019-01720 Sweden
CitationJournal: Elife / Year: 2022
Title: Protein-lipid interaction at low pH induces oligomerization of the MakA cytotoxin from .
Authors: Aftab Nadeem / Alexandra Berg / Hudson Pace / Athar Alam / Eric Toh / Jörgen Ådén / Nikola Zlatkov / Si Lhyam Myint / Karina Persson / Gerhard Gröbner / Anders Sjöstedt / Marta Bally / ...Authors: Aftab Nadeem / Alexandra Berg / Hudson Pace / Athar Alam / Eric Toh / Jörgen Ådén / Nikola Zlatkov / Si Lhyam Myint / Karina Persson / Gerhard Gröbner / Anders Sjöstedt / Marta Bally / Jonas Barandun / Bernt Eric Uhlin / Sun Nyunt Wai /
Abstract: The α-pore-forming toxins (α-PFTs) from pathogenic bacteria damage host cell membranes by pore formation. We demonstrate a remarkable, hitherto unknown mechanism by an α-PFT protein from . As part ...The α-pore-forming toxins (α-PFTs) from pathogenic bacteria damage host cell membranes by pore formation. We demonstrate a remarkable, hitherto unknown mechanism by an α-PFT protein from . As part of the MakA/B/E tripartite toxin, MakA is involved in membrane pore formation similar to other α-PFTs. In contrast, MakA in isolation induces tube-like structures in acidic endosomal compartments of epithelial cells in vitro. The present study unravels the dynamics of tubular growth, which occurs in a pH-, lipid-, and concentration-dependent manner. Within acidified organelle lumens or when incubated with cells in acidic media, MakA forms oligomers and remodels membranes into high-curvature tubes leading to loss of membrane integrity. A 3.7 Å cryo-electron microscopy structure of MakA filaments reveals a unique protein-lipid superstructure. MakA forms a pinecone-like spiral with a central cavity and a thin annular lipid bilayer embedded between the MakA transmembrane helices in its active α-PFT conformation. Our study provides insights into a novel tubulation mechanism of an α-PFT protein and a new mode of action by a secreted bacterial toxin.
History
DepositionJul 8, 2021Deposition site: PDBE / Processing site: PDBE
Revision 1.0Feb 23, 2022Provider: repository / Type: Initial release
Revision 1.1Jul 17, 2024Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / em_3d_fitting_list / em_admin / pdbx_initial_refinement_model
Item: _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id ..._em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type / _em_admin.last_update

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Structure visualization

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Assembly

Deposited unit
A: MakA tetramer
B: MakA tetramer
C: MakA tetramer
D: MakA tetramer


Theoretical massNumber of molelcules
Total (without water)156,1144
Polymers156,1144
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area8250 Å2
ΔGint-82 kcal/mol
Surface area68100 Å2

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Components

#1: Protein
MakA tetramer


Mass: 39028.582 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Vibrio cholerae O1 biovar El Tor str. N16961
Gene: VC_A0883 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q9KL64

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: HELICAL ARRAY / 3D reconstruction method: helical reconstruction

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Sample preparation

ComponentName: Vibrio cholera MakA filament / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Vibrio cholerae O1 biovar El Tor str. N16961 (bacteria)
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria)
Buffer solutionpH: 6.5
Buffer componentConc.: 120 mM / Name: Sodium Citrate
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R2/1
VitrificationInstrument: FEI VITROBOT MARK I / Cryogen name: ETHANE

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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 defocus max: 2500 nm / Nominal defocus min: 750 nm / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 43 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 QUANTUM (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 2476

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Processing

EM software
IDNameVersionCategory
1RELION3.1particle selection
2EPUimage acquisition
4CTFFIND4.1.14CTF correction
7Cootmodel fitting
10RELION3.1final Euler assignment
11RELION3.1classification
12RELION3.13D reconstruction
13PHENIX1.14-3260model refinement
CTF correctionType: NONE
Helical symmertyAngular rotation/subunit: 48.5901 ° / Axial rise/subunit: 5.84149 Å / Axial symmetry: C1
Particle selectionNum. of particles selected: 95603
3D reconstructionResolution: 3.65 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 65485 / Symmetry type: HELICAL
Atomic model buildingProtocol: OTHER / Space: REAL
Atomic model buildingPDB-ID: 6EZV

6ezv


Accession code: 6EZV / Source name: PDB / Type: experimental model

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