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- PDB-9fz5: Cryo-EM structure of LptDE-YedD complex from Escherichia Coli -

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Basic information

Entry
Database: PDB / ID: 9fz5
TitleCryo-EM structure of LptDE-YedD complex from Escherichia Coli
Components
  • LPS-assembly lipoprotein LptE
  • LPS-assembly protein LptD
  • Uncharacterized lipoprotein YedD
KeywordsMEMBRANE PROTEIN / Outer membrane protein / lipopolysaccharide transport (Lpt) complex / Lipid binding complex / LptDE Translocon / Lipocalin YedD.
Function / homology
Function and homology information


transporter complex / lipopolysaccharide transport / Gram-negative-bacterium-type cell outer membrane assembly / cell outer membrane / lipopolysaccharide binding / plasma membrane
Similarity search - Function
YedD-like protein / YedD-like superfamily / YedD-like protein / LptD, C-terminal / LPS-assembly protein LptD / : / LPS transport system D / LPS-assembly lipoprotein LptE / Lipopolysaccharide-assembly / Organic solvent tolerance-like, N-terminal ...YedD-like protein / YedD-like superfamily / YedD-like protein / LptD, C-terminal / LPS-assembly protein LptD / : / LPS transport system D / LPS-assembly lipoprotein LptE / Lipopolysaccharide-assembly / Organic solvent tolerance-like, N-terminal / LptA/(LptD N-terminal domain) LPS transport protein / Prokaryotic membrane lipoprotein lipid attachment site profile.
Similarity search - Domain/homology
LPS-assembly lipoprotein LptE / Uncharacterized lipoprotein YedD / LPS-assembly protein LptD
Similarity search - Component
Biological speciesEscherichia coli K-12 (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.57 Å
AuthorsNguyen, V.S. / Remaut, H. / Collet, J.F. / Gennaris, A. / Thouvenel, L.
Funding support Belgium, 4items
OrganizationGrant numberCountry
Fonds de la Recherche Scientifique (FNRS)WELBIO-CR-2015A-03 Belgium
Fonds de la Recherche Scientifique (FNRS)WELBIO-CR-2019C-03 Belgium
Research Foundation - Flanders (FWO)G0H5916N Belgium
Research Foundation - Flanders (FWO)12ZM421N Belgium
CitationJournal: Cell Rep / Year: 2025
Title: Optimal functioning of the Lpt bridge depends on a ternary complex between the lipocalin YedD and the LptDE translocon.
Authors: Alexandra Gennaris / Van Son Nguyen / Laurie Thouvenel / Naemi Csoma / Didier Vertommen / Bogdan Iuliu Iorga / Han Remaut / Jean-François Collet /
Abstract: The outer membrane is an efficient permeability barrier that protects gram-negative bacteria against external assaults, including many antibiotics. The unique permeability features of the outer ...The outer membrane is an efficient permeability barrier that protects gram-negative bacteria against external assaults, including many antibiotics. The unique permeability features of the outer membrane are due to the presence of lipopolysaccharide (LPS) molecules in its outer leaflet. LPS transport relies on the essential lipopolysaccharide transport (Lpt) pathway, which forms a bridge from the inner to the outer membrane. The LptDE translocon inserts LPS into the outer leaflet. Here, we identify the lipocalin YedD as a component of the translocon. Cryoelectron microscopy of the YedD-LptDE complex reveals that YedD binds LptD at a critical interface between its β-barrel and periplasmic β-taco domain. The YedD-LptDE complex is functionally relevant: under conditions where the connectivity of the β-taco and Lpt bridge is compromised, the absence of YedD decreases cell viability and causes LPS accumulation in the inner membrane. Our findings establish YedD as an Lpt component required for optimal LPS transport.
History
DepositionJul 4, 2024Deposition site: PDBE / Processing site: PDBE
Revision 1.0Apr 9, 2025Provider: repository / Type: Initial release
Revision 1.0Apr 9, 2025Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
Revision 1.0Apr 9, 2025Data content type: FSC / Data content type: FSC / Provider: repository / Type: Initial release
Revision 1.0Apr 9, 2025Data content type: Half map / Part number: 1 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Apr 9, 2025Data content type: Half map / Part number: 2 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Apr 9, 2025Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Apr 9, 2025Data content type: Mask / Part number: 1 / Data content type: Mask / Provider: repository / Type: Initial release
Revision 1.0Apr 9, 2025Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: LPS-assembly protein LptD
B: LPS-assembly lipoprotein LptE
C: Uncharacterized lipoprotein YedD


Theoretical massNumber of molelcules
Total (without water)116,8043
Polymers116,8043
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein LPS-assembly protein LptD / Organic solvent tolerance protein


Mass: 87078.438 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: lptD, imp, ostA, yabG, b0054, JW0053 / Production host: Escherichia coli (E. coli) / References: UniProt: P31554
#2: Protein LPS-assembly lipoprotein LptE / Rare lipoprotein B


Mass: 16726.266 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: lptE, rlpB, b0641, JW0636 / Production host: Escherichia coli (E. coli) / References: UniProt: P0ADC1
#3: Protein Uncharacterized lipoprotein YedD


Mass: 12999.715 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: yedD, b1928, JW1913 / Production host: Escherichia coli (E. coli) / References: UniProt: P31063
Has protein modificationY

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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: LPS-assembly complex LptDE with lipoprotein YedD / Type: COMPLEX
Details: The LptDE-YedD complex was purified from over-expression of the three components in E. coli. Strep-tag was on the C-terminal of LptE while his-tag was on the C-terminal of YedD. Double pull ...Details: The LptDE-YedD complex was purified from over-expression of the three components in E. coli. Strep-tag was on the C-terminal of LptE while his-tag was on the C-terminal of YedD. Double pull was done using 1, streptactin and 2, Ni-NTA.
Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 0.12 MDa / Experimental value: NO
Source (natural)Organism: Escherichia coli K-12 (bacteria)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 7
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMtrisaminomethaneTris1
2150 mMSodium ChlorideNaCl1
3400 mMImidazolImidazole1
40.03 %n-Dodecyl-Beta-MaltosideDDM1
SpecimenConc.: 2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: GATAN CRYOPLUNGE 3 / Cryogen name: ETHANE / Humidity: 90 % / Details: Blot 3-4 seconds

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Electron microscopy imaging

MicroscopyModel: JEOL CRYO ARM 300
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: BRIGHT FIELD / Calibrated magnification: 60000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 800 nm / Cs: 2.55 mm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: JEOL CRYOSPECPORTER
Image recordingAverage exposure time: 3 sec. / Electron dose: 60 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 13332

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Processing

EM software
IDNameVersionCategoryDetails
1crYOLO1.8.4particle selectionuse a trained model
4RELION3.1CTF correctionParticle extraction, 3D classification, and 3D reconstruction
5MotionCorr21.3CTF correctionMotion correction
6CTFFIND4CTF correctionCTF estimation
7PHENIXCTF correctionRefinement, Auto_sharpen, geometry optimisation
8Coot0.8.9CTF correctionModel building and editing
9UCSF Chimera1.15CTF correctionMap manipulation and display
10PyMOL2.2CTF correctionDisplay
11SerialEMCTF correctionImage acquisition
12cryoSPARCCTF correction2D and 3D classification and refinement
15Coot0.8.9model fitting
20cryoSPARC4.4.13D reconstructionNon-uniform refinement
27PHENIXmodel refinement
CTF correctionType: NONE
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.57 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 56171 / Symmetry type: POINT
Atomic model buildingB value: 78.86 / Protocol: OTHER / Space: REAL
Atomic model buildingSource name: AlphaFold / Type: in silico model

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