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- EMDB-27326: Structure of hemolysin A secretion system HlyB/D complex, ATP-bound -

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Basic information

Entry
Database: EMDB / ID: EMD-27326
TitleStructure of hemolysin A secretion system HlyB/D complex, ATP-bound
Map data
Sample
  • Complex: Membrane protein complex of HlyB and HlyD
    • Protein or peptide: Alpha-hemolysin translocation ATP-binding protein HlyB
    • Protein or peptide: Membrane fusion protein (MFP) family protein
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
  • Ligand: MAGNESIUM ION
Keywordshydrolase / transport / MEMBRANE PROTEIN
Function / homology
Function and homology information


type I protein secretion system complex / protein secretion by the type I secretion system / protein secretion / ABC-type transporter activity / peptidase activity / ATP hydrolysis activity / proteolysis / ATP binding / plasma membrane
Similarity search - Function
Secretion protein HlyD, conserved site / Type I secretion membrane fusion protein, HlyD family / HlyD family secretion proteins signature. / HlyD family secretion protein / : / ATPase, type I secretion system, HlyB / Peptidase C39-like A / Peptidase C39 family / Peptidase C39, bacteriocin processing / Peptidase family C39 domain profile. ...Secretion protein HlyD, conserved site / Type I secretion membrane fusion protein, HlyD family / HlyD family secretion proteins signature. / HlyD family secretion protein / : / 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 / 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: 3.4 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
DepositionJun 16, 2022-
Header (metadata) releaseSep 14, 2022-
Map releaseSep 14, 2022-
UpdateJun 12, 2024-
Current statusJun 12, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_27326.png

Downloads & links

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Map

FileDownload / File: emd_27326.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.3 Å/pix.
x 300 pix.
= 390. Å		1.3 Å/pix.
x 300 pix.
= 390. Å		1.3 Å/pix.
x 300 pix.
= 390. Å

Surface

Projections

Slices (1/3)

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Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.3 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.7
Minimum - Maximum-2.6846292 - 4.780492
Average (Standard dev.)-0.0004201567 (±0.1110659)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 390.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_27326_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_27326_half_map_2.map
Projections & Slices
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Slices (1/2)
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Histogram

Histogram (log scale)

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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 HlyD
Components
  • Complex: Membrane protein complex of HlyB and HlyD
    • Protein or peptide: Alpha-hemolysin translocation ATP-binding protein HlyB
    • Protein or peptide: Membrane fusion protein (MFP) family protein
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
  • Ligand: MAGNESIUM ION

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Supramolecule #1: Membrane protein complex of HlyB and HlyD

SupramoleculeName: Membrane protein complex of HlyB and HlyD / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Escherichia coli O6:H1 (bacteria)
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: 79.6205 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 PKILIFDQAT SALDYES EH IIMRNMHKIC KGRTVIIIAH RLSTVKNADR IIVMEKGKIV EQGKHKELLS EPESLYSYLY QLQSD

UniProtKB: Alpha-hemolysin translocation ATP-binding protein HlyB

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Macromolecule #2: Membrane fusion protein (MFP) family protein

MacromoleculeName: Membrane fusion protein (MFP) family protein / 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: 54.671672 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 RSLLHKQ AIAKHAVLEQ ENKYVEAANE LRVYKSQLEQ IESEILSAKE EYQLVTQLFK NEILDKLRQT TDNIELLTLE LEK NEERQQ ASVIRAPVSG KVQQLKVHTE GGVVTTAETL MVIVPEDDTL EVTALVQNKD IGFINVGQNA IIKVEAFPYT RYGY LVGKV KNINLDAIED QKLGLVFNVI VSVEENDLST GNKHIPLSSG MAVTAEIKTG MRSVISYLLS PLEESVTESL HER

UniProtKB: Membrane fusion protein (MFP) family protein

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Macromolecule #3: ADENOSINE-5'-TRIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 3 / Number of copies: 6 / Formula: ATP
Molecular weightTheoretical: 507.181 Da
Chemical component information

ChemComp-ATP:
ADENOSINE-5'-TRIPHOSPHATE / ATP, energy-carrying molecule*YM

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Macromolecule #4: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 6 / Formula: MG
Molecular weightTheoretical: 24.305 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

BufferpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Final reconstructionApplied symmetry - Point group: C3 (3 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 55516
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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