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- EMDB-11240: Cryo-EM structure of the E.coli HTL-BAm complex -

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Basic information

Entry
Database: EMDB / ID: EMD-11240
TitleCryo-EM structure of the E.coli HTL-BAm complex
Map data
SampleE. coli inner membrane holotranslocon in complex with the outer membrane complex BAM:
E coli HTL / E. coli BAM
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 18.23 Å
AuthorsAlvira S / Collinson S
Funding support United Kingdom, 4 items
OrganizationGrant numberCountry
Biotechnology and Biological Sciences Research Council (BBSRC)BB/S008349/1 United Kingdom
Wellcome Trust206181/Z/17/Z United Kingdom
Wellcome Trust202904/Z/16/Z United Kingdom
Biotechnology and Biological Sciences Research Council (BBSRC)BB/R000484/1 United Kingdom
CitationJournal: Elife / Year: 2020
Title: Inter-membrane association of the Sec and BAM translocons for bacterial outer-membrane biogenesis.
Authors: Sara Alvira / Daniel W Watkins / Lucy Troman / William J Allen / James S Lorriman / Gianluca Degliesposti / Eli J Cohen / Morgan Beeby / Bertram Daum / Vicki Am Gold / J Mark Skehel / Ian Collinson /
Abstract: The outer-membrane of Gram-negative bacteria is critical for surface adhesion, pathogenicity, antibiotic resistance and survival. The major constituent - hydrophobic β-barrel uter-embrane roteins ...The outer-membrane of Gram-negative bacteria is critical for surface adhesion, pathogenicity, antibiotic resistance and survival. The major constituent - hydrophobic β-barrel uter-embrane roteins (OMPs) - are first secreted across the inner-membrane through the Sec-translocon for delivery to periplasmic chaperones, for example SurA, which prevent aggregation. OMPs are then offloaded to the β-arrel ssembly achinery (BAM) in the outer-membrane for insertion and folding. We show the olo-ransocon (HTL) - an assembly of the protein-channel core-complex SecYEG, the ancillary sub-complex SecDF, and the membrane 'insertase' YidC - contacts BAM through periplasmic domains of SecDF and YidC, ensuring efficient OMP maturation. Furthermore, the proton-motive force (PMF) across the inner-membrane acts at distinct stages of protein secretion: (1) SecA-driven translocation through SecYEG and (2) communication of conformational changes via SecDF across the periplasm to BAM. The latter presumably drives efficient passage of OMPs. These interactions provide insights of inter-membrane organisation and communication, the importance of which is becoming increasingly apparent.
History
DepositionJun 29, 2020-
Header (metadata) releaseNov 18, 2020-
Map releaseNov 18, 2020-
UpdateNov 18, 2020-
Current statusNov 18, 2020Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0066
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by height
  • Surface level: 0.0066
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_11240.map.gz / Format: CCP4 / Size: 59.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.75 Å/pix.
x 250 pix.
= 437.5 Å
1.75 Å/pix.
x 250 pix.
= 437.5 Å
1.75 Å/pix.
x 250 pix.
= 437.5 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.75 Å
Density
Contour LevelBy AUTHOR: 0.0066 / Movie #1: 0.0066
Minimum - Maximum-0.0033417405 - 0.025266567
Average (Standard dev.)0.0007380842 (±0.0025354407)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions250250250
Spacing250250250
CellA=B=C: 437.5 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.751.751.75
M x/y/z250250250
origin x/y/z0.0000.0000.000
length x/y/z437.500437.500437.500
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS250250250
D min/max/mean-0.0030.0250.001

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Supplemental data

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Sample components

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Entire E. coli inner membrane holotranslocon in complex with the outer m...

EntireName: E. coli inner membrane holotranslocon in complex with the outer membrane complex BAM
Number of components: 3

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Component #1: protein, E. coli inner membrane holotranslocon in complex with th...

ProteinName: E. coli inner membrane holotranslocon in complex with the outer membrane complex BAM
Recombinant expression: No
MassTheoretical: 205 kDa
SourceSpecies: Escherichia coli (E. coli)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #2: protein, E coli HTL

ProteinName: E coli HTL / Recombinant expression: No
SourceSpecies: Escherichia coli (E. coli)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #3: protein, E. coli BAM

ProteinName: E. coli BAM / Recombinant expression: No
SourceSpecies: Escherichia coli (E. coli)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionBuffer solution: 20mM HEPES, pH 8.0 250 mM NaCl, 0.03% (w/v) DDM/0.003% (w/v) CL
pH: 8
VitrificationInstrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 90 %

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
ImagingMicroscope: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Electron dose: 1.127 e/Å2 / Illumination mode: FLOOD BEAM
LensMagnification: 79000 X (nominal) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image processing

ProcessingMethod: single particle reconstruction / Number of projections: 8448
3D reconstructionSoftware: RELION / Resolution: 18.23 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution estimation)

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