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- PDB-9krf: Alpha-hemolysin heptameric pore state bound to 10:0 PC lipid chai... -

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Basic information

Entry
Database: PDB / ID: 9krf
TitleAlpha-hemolysin heptameric pore state bound to 10:0 PC lipid chains derived from 10:0 PC liposomes
ComponentsAlpha-hemolysin
KeywordsLIPID BINDING PROTEIN / PFT / Liposomes
Function / homology
Function and homology information


cytolysis in another organism / The NLRP3 inflammasome / Purinergic signaling in leishmaniasis infection / toxin activity / extracellular region / identical protein binding
Similarity search - Function
Aerolysin/haemolysin toxin, conserved site / Aerolysin type toxins signature. / Bi-component toxin, staphylococci / Leukocidin/Hemolysin toxin / Leukocidin/Hemolysin toxin family / Leukocidin/porin MspA superfamily
Similarity search - Domain/homology
1,2-DIDECANOYL-SN-GLYCERO-3-PHOSPHOCHOLINE / Alpha-hemolysin
Similarity search - Component
Biological speciesStaphylococcus aureus (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.8 Å
AuthorsChatterjee, A. / Roy, A. / Dutta, S.
Funding support India, 1items
OrganizationGrant numberCountry
Department of Biotechnology (DBT, India) India
CitationJournal: Nat Commun / Year: 2025
Title: Structural insights into pre-pore intermediates of alpha-hemolysin in the lipidic environment.
Authors: Arnab Chatterjee / Anupam Roy / Thejas Satheesh / Partho Pratim Das / Bapan Mondal / Prithiv Kishore / Mahipal Ganji / Somnath Dutta /
Abstract: The infectious microbe Staphylococcus aureus releases an array of cytotoxic pore-forming toxins (PFTs) that severely damage the cell membrane during bacterial infection. However, the interaction ...The infectious microbe Staphylococcus aureus releases an array of cytotoxic pore-forming toxins (PFTs) that severely damage the cell membrane during bacterial infection. However, the interaction interfaces between the host cell membrane and toxin were hardly explored. So far, there are no pore, and intermediate structures of these toxins available in the presence of bio-membrane, which could elucidate the pore-forming mechanism. Here, we investigate the structure of different conformational states of this alpha-hemolysin (α-HL/Hla), a β-PFT in lipidic environment using single-particle cryo-EM. Additionally, we explore lipid destabilization by the toxin, using single-molecule imaging, confocal imaging, and validation of lipid-protein interactions using mutational studies. We elucidate eight cryo-EM structures of wildtype α-HL with various liposomes, which are composed of either 10:0 PC or Egg-PC/Cholesterol or Egg-PC/Sphingomyelin or 10:0 PC/Sphingomyelin. Our structural and biophysical studies confirm that different chain lengths and various membrane compositions facilitate the formation of intermediate pre-pores and complete pore species. We also demonstrate that the percentage of pre-pore population increases due to sphingomyelin-induced membrane rigidity. Altogether, this study unveils the structure-function analysis of the pre-pore to pore transition of wildtype α-HL during its crosstalk with the lipid membrane.
History
DepositionNov 27, 2024Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jun 11, 2025Provider: repository / Type: Initial release
Revision 1.1Jul 23, 2025Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
G: Alpha-hemolysin
A: Alpha-hemolysin
B: Alpha-hemolysin
C: Alpha-hemolysin
D: Alpha-hemolysin
E: Alpha-hemolysin
F: Alpha-hemolysin
hetero molecules


Theoretical massNumber of molelcules
Total (without water)236,99714
Polymers233,0307
Non-polymers3,9677
Water00
1


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

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Components

#1: Protein
Alpha-hemolysin / Alpha-HL / Alpha-toxin


Mass: 33290.000 Da / Num. of mol.: 7
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Staphylococcus aureus (bacteria) / Gene: hly, hla / Production host: Escherichia coli (E. coli) / References: UniProt: P09616
#2: Chemical
ChemComp-P1O / 1,2-DIDECANOYL-SN-GLYCERO-3-PHOSPHOCHOLINE


Mass: 566.728 Da / Num. of mol.: 7 / Source method: obtained synthetically / Formula: C28H57NO8P / Feature type: SUBJECT OF INVESTIGATION / Comment: DDPC, phospholipid*YM
Has ligand of interestY
Has protein modificationN

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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: Alpha-hemolysin heptameric pore state bound to 10:0 PC lipid chains derived from 10:0 PC liposomes
Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Source (natural)Organism: Staphylococcus aureus (bacteria)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: OTHER
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 3500 nm / Nominal defocus min: 750 nm
Image recordingElectron dose: 40 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

EM softwareName: PHENIX / Category: model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 127106 / Symmetry type: POINT
RefinementHighest resolution: 2.8 Å
Stereochemistry target values: REAL-SPACE (WEIGHTED MAP SUM AT ATOM CENTERS)
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00517108
ELECTRON MICROSCOPYf_angle_d0.45523128
ELECTRON MICROSCOPYf_dihedral_angle_d12.5316440
ELECTRON MICROSCOPYf_chiral_restr0.0422443
ELECTRON MICROSCOPYf_plane_restr0.0032961

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