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- EMDB-25116: Structure of hemolysin A secretion system HlyB/D complex -

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Basic information

Entry
Database: EMDB / ID: EMD-25116
TitleStructure of hemolysin A secretion system HlyB/D complex
Map data
Sample
  • Complex: Membrane protein complex of HlyB and HlyDBiological membrane
    • Protein or peptide: Alpha-hemolysin translocation ATP-binding protein HlyB
  • Protein or peptide: Membrane fusion protein (MFP) family protein,Hemolysin secretion protein D, chromosomalLipid bilayer fusion
  • Ligand: [(2~{R})-1-[2-azanylethoxy(oxidanyl)phosphoryl]oxy-3-hexadecanoyloxy-propan-2-yl] (~{Z})-octadec-9-enoate
Function / homology
Function and homology information


protein secretion by the type I secretion system / type I protein secretion system complex / : / toxin transmembrane transporter activity / secretion by cell / protein secretion / ABC-type transporter activity / peptidase activity / killing of cells of another organism / membrane => GO:0016020 ...protein secretion by the type I secretion system / type I protein secretion system complex / : / toxin transmembrane transporter activity / secretion by cell / protein secretion / ABC-type transporter activity / peptidase activity / killing of cells of another organism / membrane => GO:0016020 / proteolysis / ATP binding / plasma membrane
Similarity search - Function
Secretion protein HlyD, conserved site / Type I secretion membrane fusion protein, HlyD family / HlyD family secretion protein / HlyD family secretion proteins signature. / ATPase, type I secretion system, HlyB / Peptidase C39-like A / Peptidase C39 family / Peptidase C39, bacteriocin processing / Peptidase family C39 domain profile. / Type 1 protein exporter ...Secretion protein HlyD, conserved site / Type I secretion membrane fusion protein, HlyD family / HlyD family secretion protein / HlyD family secretion proteins signature. / ATPase, type I secretion system, HlyB / Peptidase C39-like A / Peptidase C39 family / Peptidase C39, bacteriocin processing / Peptidase family C39 domain profile. / Type 1 protein exporter / ABC transporter transmembrane region / ABC transporter type 1, transmembrane domain / ABC transporter integral membrane type-1 fused domain profile. / ABC transporter type 1, transmembrane domain superfamily / ABC transporter-like, conserved site / ABC transporters family signature. / ABC transporter / ABC transporter-like, ATP-binding domain / ATP-binding cassette, ABC transporter-type domain profile. / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Membrane fusion protein (MFP) family protein / Hemolysin secretion protein D, chromosomal / Alpha-hemolysin translocation ATP-binding protein HlyB
Similarity search - Component
Biological speciesEscherichia coli O6:H1 (bacteria) / Escherichia coli CFT073 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsZhao H / Chen J
Funding support France, United States, 2 items
OrganizationGrant numberCountry
Human Frontier Science Program (HFSP) France
Howard Hughes Medical Institute (HHMI) United States
CitationJournal: Cell / Year: 2022
Title: The hemolysin A secretion system is a multi-engine pump containing three ABC transporters.
Authors: Hongtu Zhao / James Lee / Jue Chen /
Abstract: Type 1 secretion systems (T1SSs) are widespread in pathogenic Gram-negative bacteria, extruding protein substrates following synthesis of the entire polypeptide. The Escherichia coli hemolysin A ...Type 1 secretion systems (T1SSs) are widespread in pathogenic Gram-negative bacteria, extruding protein substrates following synthesis of the entire polypeptide. The Escherichia coli hemolysin A secretion system has long been considered a prototype in structural and mechanistic studies of T1SSs. Three membrane proteins-an inner membrane ABC transporter HlyB, an adaptor protein HlyD, and an outer membrane porin TolC-are required for secretion. However, the stoichiometry and structure of the complex are unknown. Here, cryo-electron microscopy (cryo-EM) structures determined in two conformations reveal that the inner membrane complex is a hetero-dodecameric assembly comprising three HlyB homodimers and six HlyD subunits. Functional studies indicate that oligomerization of HlyB and HlyD is essential for protein secretion and that polypeptides translocate through a canonical ABC transporter pathway in HlyB. Our data suggest that T1SSs entail three ABC transporters, one that functions as a protein channel and two that allosterically power the translocation process.
History
DepositionOct 7, 2021-
Header (metadata) releaseSep 14, 2022-
Map releaseSep 14, 2022-
UpdateSep 14, 2022-
Current statusSep 14, 2022Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_25116.png

Downloads & links

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Map

FileDownload / File: emd_25116.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.07 Å
Density
Contour LevelBy AUTHOR: 0.4
Minimum - Maximum-2.3838627 - 4.4336143
Average (Standard dev.)-0.0011432696 (±0.10447862)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 410.88 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : Membrane protein complex of HlyB and HlyD

EntireName: Membrane protein complex of HlyB and HlyDBiological membrane
Components
  • Complex: Membrane protein complex of HlyB and HlyDBiological membrane
    • Protein or peptide: Alpha-hemolysin translocation ATP-binding protein HlyB
  • Protein or peptide: Membrane fusion protein (MFP) family protein,Hemolysin secretion protein D, chromosomalLipid bilayer fusion
  • Ligand: [(2~{R})-1-[2-azanylethoxy(oxidanyl)phosphoryl]oxy-3-hexadecanoyloxy-propan-2-yl] (~{Z})-octadec-9-enoate

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Supramolecule #1: Membrane protein complex of HlyB and HlyD

SupramoleculeName: Membrane protein complex of HlyB and HlyD / type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Escherichia coli O6:H1 (bacteria)
Recombinant expressionOrganism: Escherichia coli (E. coli)
Molecular weightTheoretical: 810 KDa

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Macromolecule #1: Alpha-hemolysin translocation ATP-binding protein HlyB

MacromoleculeName: Alpha-hemolysin translocation ATP-binding protein HlyB
type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli CFT073 (bacteria) / Strain: CFT073 / ATCC 700928 / UPEC
Molecular weightTheoretical: 79.621492 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MDSCHKIDYG LYALEILAQY HNVSVNPEEI KHRFDTDGTG LGLTSWLLAA KSLELKVKQV KKTIDRLNFI SLPALVWRED GRHFILTKV SKEANRYLIF DLEQRNPRVL EQSEFEALYQ GHIILIASRS SVTGKLAKFD FTWFIPAIIK YRKIFIETLV V SVFLQLFA ...String:
MDSCHKIDYG LYALEILAQY HNVSVNPEEI KHRFDTDGTG LGLTSWLLAA KSLELKVKQV KKTIDRLNFI SLPALVWRED GRHFILTKV SKEANRYLIF DLEQRNPRVL EQSEFEALYQ GHIILIASRS SVTGKLAKFD FTWFIPAIIK YRKIFIETLV V SVFLQLFA LITPLFFQVV MDKVLVHRGF STLNVITVAL SVVVVFEIIL SGLRTYIFAH STSRIDVELG AKLFRHLLAL PI SYFESRR VGDTVARVRE LDQIRNFLTG QALTSVLDLL FSFIFFAVMW YYSPKLTLVI LFSLPCYAAW SVFISPILRR RLD DKFSRN ADNQSFLVES VTAINTIKAM AVSPQMTNIW DKQLAGYVAA GFKVTVLATI GQQGIQLIQK TVMIINLWLG AHLV ISGDL SIGQLIAFNM LAGQIVAPVI RLAQIWQDFQ QVGISVTRLG DVLNSPTESY HGKLALPEIN GNITFRNIRF RYKPD SPVI LDNINLSIKQ GEVIGIVGRS GSGKSTLTKL IQRFYIPENG QVLIDGHDLA LADPNWLRRQ VGVVLQDNVL LNRSII DNI SLANPGMSVE KVIYAAKLAG AHDFISELRE GYNTIVGEQG AGLSGGQRQR IAIARALVNN PKILIFDEAT SALDYES EH IIMRNMHKIC KGRTVIIIAH RLSTVKNADR IIVMEKGKIV EQGKHKELLS EPESLYSYLY QLQSD

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Macromolecule #2: Membrane fusion protein (MFP) family protein,Hemolysin secretion ...

MacromoleculeName: Membrane fusion protein (MFP) family protein,Hemolysin secretion protein D, chromosomal
type: protein_or_peptide / ID: 2 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli CFT073 (bacteria) / Strain: CFT073 / ATCC 700928 / UPEC
Molecular weightTheoretical: 40.748895 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MKTWLMGFSE FLLRYKLVWS ETWKIRKQLD TPVREKDENE FLPAHLELIE TPVSRRPRLV AYFIMGFLVI AVILSVLGQV EIVATANGK LTLSGRSKEI KPIENSIVKE IIVKEGESVR KGDVLLKLTA LGAEADTLKT QSSLLQTRLE QTRYQILSRS I ELNKLPEL ...String:
MKTWLMGFSE FLLRYKLVWS ETWKIRKQLD TPVREKDENE FLPAHLELIE TPVSRRPRLV AYFIMGFLVI AVILSVLGQV EIVATANGK LTLSGRSKEI KPIENSIVKE IIVKEGESVR KGDVLLKLTA LGAEADTLKT QSSLLQTRLE QTRYQILSRS I ELNKLPEL KLPDEPYFQN VSEEEVLRLT SLIKEQFSTW QNQKYQKELN LDKKRAERLT ILARINRYEN LSRVEKSRLD DF DDTLEVT ALVQNKDIGF INVGQNAIIK VEAFPYTRYG YLVGKVKNIN LDAIEDQKLG LVFNVIVSVE ENDLSTGNKH IPL SSGMAV TAEIKTGMRS VISYLLSPLE ESVTESLHER

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Macromolecule #3: [(2~{R})-1-[2-azanylethoxy(oxidanyl)phosphoryl]oxy-3-hexadecanoyl...

MacromoleculeName: [(2~{R})-1-[2-azanylethoxy(oxidanyl)phosphoryl]oxy-3-hexadecanoyloxy-propan-2-yl] (~{Z})-octadec-9-enoate
type: ligand / ID: 3 / Number of copies: 37 / Formula: 6OU
Molecular weightTheoretical: 717.996 Da
Chemical component information

ChemComp-6OU:
[(2~{R})-1-[2-azanylethoxy(oxidanyl)phosphoryl]oxy-3-hexadecanoyloxy-propan-2-yl] (~{Z})-octadec-9-enoate / phospholipid*YM

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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

BufferpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.0 µm
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 70.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 136123

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