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- EMDB-12950: Cryo-EM structure of undecameric SlyB from Escherichia coli K12 -

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Basic information

Entry
Database: EMDB / ID: EMD-12950
TitleCryo-EM structure of undecameric SlyB from Escherichia coli K12
Map data
Sample
  • Complex: Outer membrane lipoprotein SlyB
    • Protein or peptide: Outer membrane lipoprotein slyB
  • Ligand: PALMITIC ACID
  • Ligand: GLYCEROL
  • Ligand: (2~{R},4~{R},5~{R},6~{R})-6-[(1~{R})-1,2-bis(oxidanyl)ethyl]-2-[[(2~{R},3~{S},4~{R},5~{R},6~{R})-5-[[(~{E},3~{R})-3-dodecanoyloxytetradec-5-enoyl]amino]-6-[[(2~{R},3~{S},4~{R},5~{R},6~{R})-3-oxidanyl-5-[[(~{E},3~{R})-3-oxidanyltetradec-11-enoyl]amino]-4-[(~{E},3~{R})-3-oxidanyltetradec-5-enoyl]oxy-6-phosphonooxy-oxan-2-yl]methoxy]-3-phosphonooxy-4-[(~{E},3~{R})-3-tetradecanoyloxytetradec-7-enoyl]oxy-oxan-2-yl]methoxy]-4,5-bis(oxidanyl)oxane-2-carboxylic acid
  • Ligand: 2-(HEXADECANOYLOXY)-1-[(PHOSPHONOOXY)METHYL]ETHYL HEXADECANOATE
KeywordsOUTER MEMBRANE CHAPERONE / 2TM GLYCINE ZIPPER / OUTER MEMBRANE LIPOPROTEIN SLYB / LPS-LP BINDING PROTEIN / MEMBRANE PROTEIN
Function / homologyGlycine zipper 2TM domain / Glycine zipper 2TM domain / cell outer membrane / Prokaryotic membrane lipoprotein lipid attachment site profile. / Outer membrane lipoprotein slyB
Function and homology information
Biological speciesEscherichia coli BW25113 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsNguyen VS / Remaut H
Funding support Belgium, 4 items
OrganizationGrant numberCountry
Research Foundation - Flanders (FWO)G0G0818N Belgium
Research Foundation - Flanders (FWO)G0H5916N Belgium
Research Foundation - Flanders (FWO)FWOTM903 Belgium
Research Foundation - Flanders (FWO)12ZM421N Belgium
CitationJournal: Nature / Year: 2023
Title: SlyB encapsulates outer membrane proteins in stress-induced lipid nanodomains
Authors: Janssens A / Nguyen VS / Cecil AJ / Van der Verren SE / Timmerman E / Deghelt M / Pak AJ / Collet JF / Impens F / Remaut H
History
DepositionMay 14, 2021-
Header (metadata) releaseAug 3, 2022-
Map releaseAug 3, 2022-
UpdateDec 20, 2023-
Current statusDec 20, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_12950.png

Downloads & links

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Map

FileDownload / File: emd_12950.map.gz / Format: CCP4 / Size: 91.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 0.784 Å
Density
Contour LevelBy AUTHOR: 0.0085
Minimum - Maximum-0.025585115 - 0.04726879
Average (Standard dev.)0.00013731938 (±0.0019468331)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions288288288
Spacing288288288
CellA=B=C: 225.79199 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : Outer membrane lipoprotein SlyB

EntireName: Outer membrane lipoprotein SlyB
Components
  • Complex: Outer membrane lipoprotein SlyB
    • Protein or peptide: Outer membrane lipoprotein slyB
  • Ligand: PALMITIC ACID
  • Ligand: GLYCEROL
  • Ligand: (2~{R},4~{R},5~{R},6~{R})-6-[(1~{R})-1,2-bis(oxidanyl)ethyl]-2-[[(2~{R},3~{S},4~{R},5~{R},6~{R})-5-[[(~{E},3~{R})-3-dodecanoyloxytetradec-5-enoyl]amino]-6-[[(2~{R},3~{S},4~{R},5~{R},6~{R})-3-oxidanyl-5-[[(~{E},3~{R})-3-oxidanyltetradec-11-enoyl]amino]-4-[(~{E},3~{R})-3-oxidanyltetradec-5-enoyl]oxy-6-phosphonooxy-oxan-2-yl]methoxy]-3-phosphonooxy-4-[(~{E},3~{R})-3-tetradecanoyloxytetradec-7-enoyl]oxy-oxan-2-yl]methoxy]-4,5-bis(oxidanyl)oxane-2-carboxylic acid
  • Ligand: 2-(HEXADECANOYLOXY)-1-[(PHOSPHONOOXY)METHYL]ETHYL HEXADECANOATE

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Supramolecule #1: Outer membrane lipoprotein SlyB

SupramoleculeName: Outer membrane lipoprotein SlyB / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: Undecameric SlyB was purified from overexpression of SlyB-TEV-HIS in E. coli using using 2-step Ni-IMAC purification.
Source (natural)Organism: Escherichia coli BW25113 (bacteria) / Location in cell: Outer membrane
Molecular weightTheoretical: 200 KDa

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Macromolecule #1: Outer membrane lipoprotein slyB

MacromoleculeName: Outer membrane lipoprotein slyB / type: protein_or_peptide / ID: 1 / Number of copies: 11 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli BW25113 (bacteria) / Strain: 2002017
Molecular weightTheoretical: 15.613606 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MIKRVLVVSM VGLSLVGCVN NDTLSGDVYT ASEAKQVQNV SYGTIVNVRP VQIQGGDDSN VIGAIGGAVL GGFLGNTVGG GTGRSLATA AGAVAGGVAG QGVQSAMNKT QGVELEIRKD DGNTIMVVQK QGNTRFSPGQ RVVLASNGSQ VTVSPR

UniProtKB: Outer membrane lipoprotein slyB

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Macromolecule #2: PALMITIC ACID

MacromoleculeName: PALMITIC ACID / type: ligand / ID: 2 / Number of copies: 33 / Formula: PLM
Molecular weightTheoretical: 256.424 Da
Chemical component information

ChemComp-PLM:
PALMITIC ACID / Palmitic acid

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Macromolecule #3: GLYCEROL

MacromoleculeName: GLYCEROL / type: ligand / ID: 3 / Number of copies: 11 / Formula: GOL
Molecular weightTheoretical: 92.094 Da
Chemical component information

ChemComp-GOL:
GLYCEROL / Glycerol

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Macromolecule #4: (2~{R},4~{R},5~{R},6~{R})-6-[(1~{R})-1,2-bis(oxidanyl)ethyl]-2-[[...

MacromoleculeName: (2~{R},4~{R},5~{R},6~{R})-6-[(1~{R})-1,2-bis(oxidanyl)ethyl]-2-[[(2~{R},3~{S},4~{R},5~{R},6~{R})-5-[[(~{E},3~{R})-3-dodecanoyloxytetradec-5-enoyl]amino]-6-[[(2~{R},3~{S},4~{R},5~{R},6~{R})-3- ...Name: (2~{R},4~{R},5~{R},6~{R})-6-[(1~{R})-1,2-bis(oxidanyl)ethyl]-2-[[(2~{R},3~{S},4~{R},5~{R},6~{R})-5-[[(~{E},3~{R})-3-dodecanoyloxytetradec-5-enoyl]amino]-6-[[(2~{R},3~{S},4~{R},5~{R},6~{R})-3-oxidanyl-5-[[(~{E},3~{R})-3-oxidanyltetradec-11-enoyl]amino]-4-[(~{E},3~{R})-3-oxidanyltetradec-5-enoyl]oxy-6-phosphonooxy-oxan-2-yl]methoxy]-3-phosphonooxy-4-[(~{E},3~{R})-3-tetradecanoyloxytetradec-7-enoyl]oxy-oxan-2-yl]methoxy]-4,5-bis(oxidanyl)oxane-2-carboxylic acid
type: ligand / ID: 4 / Number of copies: 11 / Formula: L8Z
Molecular weightTheoretical: 2.010478 KDa
Chemical component information

ChemComp-L8Z:
(2~{R},4~{R},5~{R},6~{R})-6-[(1~{R})-1,2-bis(oxidanyl)ethyl]-2-[[(2~{R},3~{S},4~{R},5~{R},6~{R})-5-[[(~{E},3~{R})-3-dodecanoyloxytetradec-5-enoyl]amino]-6-[[(2~{R},3~{S},4~{R},5~{R},6~{R})-3-oxidanyl-5-[[(~{E},3~{R})-3-oxidanyltetradec-11-enoyl]amino]-4-[(~{E},3~{R})-3-oxidanyltetradec-5-enoyl]oxy-6-phosphonooxy-oxan-2-yl]methoxy]-3-phosphonooxy-4-[(~{E},3~{R})-3-tetradecanoyloxytetradec-7-enoyl]oxy-oxan-2-yl]methoxy]-4,5-bis(oxidanyl)oxane-2-carboxylic acid

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Macromolecule #5: 2-(HEXADECANOYLOXY)-1-[(PHOSPHONOOXY)METHYL]ETHYL HEXADECANOATE

MacromoleculeName: 2-(HEXADECANOYLOXY)-1-[(PHOSPHONOOXY)METHYL]ETHYL HEXADECANOATE
type: ligand / ID: 5 / Number of copies: 11 / Formula: LPP
Molecular weightTheoretical: 648.891 Da
Chemical component information

ChemComp-LPP:
2-(HEXADECANOYLOXY)-1-[(PHOSPHONOOXY)METHYL]ETHYL HEXADECANOATE / phospholipid*YM

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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

Concentration0.04 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
150.0 mMNaClSodium chloridesodium chloride
20.0 mMTristris(hydroxymethyl)aminomethane
0.03 %DDMN-Dodecyl-beta-Maltoside
GridModel: Quantifoil R2/1 / Material: COPPER / Support film - Material: GRAPHENE OXIDE / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Instrument: GATAN CRYOPLUNGE 3 / Details: Back-blotting for 4 seconds before plunging.

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Electron microscopy

MicroscopeJEOL CRYO ARM 300
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.55 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.6 µm / Nominal magnification: 60000
Sample stageSpecimen holder model: JEOL CRYOSPECPORTER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 6352 / Average exposure time: 3.0 sec. / Average electron dose: 61.0 e/Å2

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Image processing

Startup modelType of model: NONE / Details: AB INITIO MODEL
Initial angle assignmentType: NOT APPLICABLE
Final 3D classificationSoftware - Name: RELION (ver. 3.1)
Final angle assignmentType: NOT APPLICABLE
Final reconstructionApplied symmetry - Point group: C11 (11 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 145472
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL / Overall B value: 35
Output model

PDB-7ojg:
CRYO-EM STRUCTURE OF UNDECAMERIC SLYB FROM ESCHERICHIA COLI K12

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