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- PDB-6v0d: Lipophilic Envelope-spanning Tunnel B (LetB), Model 2 -

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Basic information

Entry
Database: PDB / ID: 6v0d
TitleLipophilic Envelope-spanning Tunnel B (LetB), Model 2
ComponentsIntermembrane transport protein YebT
KeywordsLIPID TRANSPORT / bacterial cell envelope / MCE / YebT
Function / homology: / Mce/MlaD / MlaD protein / intermembrane lipid transfer / outer membrane-bounded periplasmic space / identical protein binding / plasma membrane / Intermembrane transport protein YebT
Function and homology information
Biological speciesEscherichia coli (E. coli)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.49 Å
AuthorsIsom, G.L. / Coudray, N. / MacRae, M.R. / McManus, C.T. / Ekiert, D.C. / Bhabha, G.
Funding support United States, 5items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R00GM112982 United States
Other privateDFS-20-16 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM128777-01 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)ACB-12002 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)AGM-12006 United States
CitationJournal: Cell / Year: 2020
Title: LetB Structure Reveals a Tunnel for Lipid Transport across the Bacterial Envelope.
Authors: Georgia L Isom / Nicolas Coudray / Mark R MacRae / Collin T McManus / Damian C Ekiert / Gira Bhabha /
Abstract: Gram-negative bacteria are surrounded by an outer membrane composed of phospholipids and lipopolysaccharide, which acts as a barrier and contributes to antibiotic resistance. The systems that mediate ...Gram-negative bacteria are surrounded by an outer membrane composed of phospholipids and lipopolysaccharide, which acts as a barrier and contributes to antibiotic resistance. The systems that mediate phospholipid trafficking across the periplasm, such as MCE (Mammalian Cell Entry) transporters, have not been well characterized. Our ~3.5 Å cryo-EM structure of the E. coli MCE protein LetB reveals an ~0.6 megadalton complex that consists of seven stacked rings, with a central hydrophobic tunnel sufficiently long to span the periplasm. Lipids bind inside the tunnel, suggesting that it functions as a pathway for lipid transport. Cryo-EM structures in the open and closed states reveal a dynamic tunnel lining, with implications for gating or substrate translocation. Our results support a model in which LetB establishes a physical link between the two membranes and creates a hydrophobic pathway for the translocation of lipids across the periplasm.
History
DepositionNov 18, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0May 6, 2020Provider: repository / Type: Initial release
Revision 1.1Nov 25, 2020Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID
Revision 1.2Mar 6, 2024Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / em_3d_fitting_list / pdbx_initial_refinement_model
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type

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Structure visualization

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Assembly

Deposited unit
A: Intermembrane transport protein YebT
B: Intermembrane transport protein YebT
C: Intermembrane transport protein YebT
D: Intermembrane transport protein YebT
E: Intermembrane transport protein YebT
F: Intermembrane transport protein YebT


Theoretical massNumber of molelcules
Total (without water)538,4646
Polymers538,4646
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Intermembrane transport protein YebT


Mass: 89744.031 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (strain K12) (bacteria)
Strain: K12 / Gene: yebT, b1834, JW1823 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P76272

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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: Lipophilic Envelope-spanning Tunnel B (LetB), Map 2, Model 2
Type: COMPLEX / Details: Cryo-EM reconstruction of full map, class 1.3 / Entity ID: all / Source: NATURAL
Molecular weightValue: 0.56 MDa / Experimental value: NO
Source (natural)Organism: Escherichia coli K-12 (bacteria)
Buffer solutionpH: 8
SpecimenConc.: 0.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK III / Cryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Cs: 2.7 mm
Specimen holderSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 80 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 4906
Image scansMovie frames/image: 50 / Used frames/image: 1-50

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Processing

EM software
IDNameVersionCategory
1RELION2.1particle selection
2SerialEMimage acquisition
4GctfCTF correction
7Cootmodel fitting
9PHENIXmodel refinement
10RELION2.1initial Euler assignment
11RELION2.1final Euler assignment
12RELION2.1classification
13RELION2.13D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 731231
SymmetryPoint symmetry: C6 (6 fold cyclic)
3D reconstructionResolution: 3.49 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 57157 / Symmetry type: POINT
Atomic model buildingPDB-ID: 5UW2
Pdb chain-ID: A / Accession code: 5UW2 / Source name: PDB / Type: experimental model

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