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- EMDB-12256: Structure of the autoinducer-2 exporter TqsA from E. coli -

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Basic information

Entry
Database: EMDB / ID: EMD-12256
TitleStructure of the autoinducer-2 exporter TqsA from E. coli
Map dataC5 symmetry imposed final map for TqsA
Sample
  • Complex: Pentameric TqsA
    • Protein or peptide: AI-2 transport protein TqsA
Function / homologyautoinducer AI-2 transmembrane transport / Transmembrane protein TqsA-like / AI-2E family transporter / quorum sensing / efflux transmembrane transporter activity / plasma membrane => GO:0005886 / transmembrane transport / AI-2 transport protein TqsA
Function and homology information
Biological speciesEscherichia coli (E. coli) / Escherichia coli (strain K12) (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsKhera R / Xie H / Michel H
Funding support Germany, 1 items
OrganizationGrant numberCountry
Max Planck Society Germany
CitationJournal: EMBO J / Year: 2022
Title: Cryo-EM structures of pentameric autoinducer-2 exporter from Escherichia coli reveal its transport mechanism.
Authors: Radhika Khera / Ahmad R Mehdipour / Jani R Bolla / Joerg Kahnt / Sonja Welsch / Ulrich Ermler / Cornelia Muenke / Carol V Robinson / Gerhard Hummer / Hao Xie / Hartmut Michel /
Abstract: Bacteria utilize small extracellular molecules to communicate in order to collectively coordinate their behaviors in response to the population density. Autoinducer-2 (AI-2), a universal molecule for ...Bacteria utilize small extracellular molecules to communicate in order to collectively coordinate their behaviors in response to the population density. Autoinducer-2 (AI-2), a universal molecule for both intra- and inter-species communication, is involved in the regulation of biofilm formation, virulence, motility, chemotaxis, and antibiotic resistance. While many studies have been devoted to understanding the biosynthesis and sensing of AI-2, very little information is available on its export. The protein TqsA from Escherichia coli, which belongs to the AI-2 exporter superfamily, has been shown to export AI-2. Here, we report the cryogenic electron microscopic structures of two AI-2 exporters (TqsA and YdiK) from E. coli at 3.35 Å and 2.80 Å resolutions, respectively. Our structures suggest that the AI-2 exporter exists as a homo-pentameric complex. In silico molecular docking and native mass spectrometry experiments were employed to demonstrate the interaction between AI-2 and TqsA, and the results highlight the functional importance of two helical hairpins in substrate binding. We propose that each monomer works as an independent functional unit utilizing an elevator-type transport mechanism.
History
DepositionJan 25, 2021-
Header (metadata) releaseMay 11, 2022-
Map releaseMay 11, 2022-
UpdateSep 28, 2022-
Current statusSep 28, 2022Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_12256.png

Downloads & links

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Map

FileDownload / File: emd_12256.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationC5 symmetry imposed final map for TqsA
Voxel sizeX=Y=Z: 0.837 Å
Density
Contour LevelBy AUTHOR: 0.0109
Minimum - Maximum-0.024183096 - 0.06282771
Average (Standard dev.)0.00029979044 (±0.0017810285)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 267.84 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_12256_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: C1 symmetry map

Fileemd_12256_additional_1.map
AnnotationC1 symmetry map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map2

Fileemd_12256_half_map_1.map
AnnotationHalf map2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map1

Fileemd_12256_half_map_2.map
AnnotationHalf map1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Pentameric TqsA

EntireName: Pentameric TqsA
Components
  • Complex: Pentameric TqsA
    • Protein or peptide: AI-2 transport protein TqsA

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Supramolecule #1: Pentameric TqsA

SupramoleculeName: Pentameric TqsA / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Escherichia coli (E. coli)
Recombinant expressionOrganism: Escherichia coli (E. coli) / Recombinant plasmid: pBADC3
Molecular weightExperimental: 187 KDa

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Macromolecule #1: AI-2 transport protein TqsA

MacromoleculeName: AI-2 transport protein TqsA / type: protein_or_peptide / ID: 1 / Number of copies: 5 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli (strain K12) (bacteria) / Strain: K12
Molecular weightTheoretical: 37.563543 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MAKPIITLNG LKIVIMLGML VIILCGIRFA AEIIVPFILA LFIAVILNPL VQHMVRWRVP RVLAVSILMT IIVMAMVLLL AYLGSALNE LTRTLPQYRN SIMTPLQALE PLLQRVGIDV SVDQLAHYID PNAAMTLLTN LLTQLSNAMS SIFLLLLTVL F MLLEVPQL ...String:
MAKPIITLNG LKIVIMLGML VIILCGIRFA AEIIVPFILA LFIAVILNPL VQHMVRWRVP RVLAVSILMT IIVMAMVLLL AYLGSALNE LTRTLPQYRN SIMTPLQALE PLLQRVGIDV SVDQLAHYID PNAAMTLLTN LLTQLSNAMS SIFLLLLTVL F MLLEVPQL PGKFQQMMAR PVEGMAAIQR AIDSVSHYLV LKTAISIITG LVAWAMLAAL DVRFAFVWGL LAFALNYIPN IG SVLAAIP PIAQVLVFNG FYEALLVLAG YLLINLVFGN ILEPRIMGRG LGLSTLVVFL SLIFWGWLLG PVGMLLSVPL TII VKIALE QTAGGQSIAV LLSDLNKE

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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

Concentration3.4 mg/mL
BufferpH: 7.5 / Details: 50 mM Tris (pH 7.5), 150 mM NaCl and 0.006% GDN
GridModel: C-flat-1.2/1.3 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK III / Details: blot time 4 s, blot force +20.
DetailsIts a membrane protein and for purification, glyco-diosgenin was used.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: OTHER / Cs: 2.7 mm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number grids imaged: 1 / Number real images: 5452 / Average electron dose: 80.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1218552
CTF correctionSoftware - Name: CTFFIND (ver. 4.0)
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final 3D classificationNumber classes: 4 / Software - Name: RELION (ver. 3.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C5 (5 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 141366
FSC plot (resolution estimation)

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