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- EMDB-25731: CryoEM map of the YejM/LapB complex with periplasmic domain -

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Basic information

Entry
Database: EMDB / ID: EMD-25731
TitleCryoEM map of the YejM/LapB complex with periplasmic domain
Map datasharpened map with a B factor of -216 A2
Sample
  • Complex: complex of YejM and YciM
    • Protein or peptide: YciM
    • Protein or peptide: YejM
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.1 Å
AuthorsMi W / Shu S
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID) United States
CitationJournal: Nat Commun / Year: 2022
Title: Regulatory mechanisms of lipopolysaccharide synthesis in Escherichia coli.
Authors: Sheng Shu / Wei Mi /
Abstract: Lipopolysaccharide (LPS) is an essential glycolipid and forms a protective permeability barrier for most Gram-negative bacteria. In E. coli, LPS levels are under feedback control, achieved by FtsH- ...Lipopolysaccharide (LPS) is an essential glycolipid and forms a protective permeability barrier for most Gram-negative bacteria. In E. coli, LPS levels are under feedback control, achieved by FtsH-mediated degradation of LpxC, which catalyzes the first committed step in LPS synthesis. FtsH is a membrane-bound AAA+ protease, and its protease activity toward LpxC is regulated by essential membrane proteins LapB and YejM. However, the regulatory mechanisms are elusive. We establish an in vitro assay to analyze the kinetics of LpxC degradation and demonstrate that LapB is an adaptor protein that utilizes its transmembrane helix to interact with FtsH and its cytoplasmic domains to recruit LpxC. Our YejM/LapB complex structure reveals that YejM is an anti-adaptor protein, competing with FtsH for LapB to inhibit LpxC degradation. Structural analysis unravels that LapB and LPS have overlapping binding sites in YejM. Thus, LPS levels control formation of the YejM/LapB complex to determine LpxC protein levels.
History
DepositionDec 14, 2021-
Header (metadata) releaseAug 17, 2022-
Map releaseAug 17, 2022-
UpdateAug 17, 2022-
Current statusAug 17, 2022Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_25731.png

Downloads & links

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Map

FileDownload / File: emd_25731.map.gz / Format: CCP4 / Size: 27 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationsharpened map with a B factor of -216 A2
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.35 Å/pix.
x 192 pix.
= 258.432 Å		1.35 Å/pix.
x 192 pix.
= 258.432 Å		1.35 Å/pix.
x 192 pix.
= 258.432 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.346 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.227
Minimum - Maximum-0.9974693 - 1.5517206
Average (Standard dev.)0.007672193 (±0.044256203)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions192192192
Spacing192192192
CellA=B=C: 258.432 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: original reconstruction without sharpening

Fileemd_25731_additional_1.map
Annotationoriginal reconstruction without sharpening
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_25731_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_25731_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : complex of YejM and YciM

EntireName: complex of YejM and YciM
Components
  • Complex: complex of YejM and YciM
    • Protein or peptide: YciM
    • Protein or peptide: YejM

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Supramolecule #1: complex of YejM and YciM

SupramoleculeName: complex of YejM and YciM / type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Escherichia coli (E. coli)
Recombinant expressionOrganism: Escherichia coli (E. coli)
Molecular weightTheoretical: 225.556 kDa/nm

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Macromolecule #1: YciM

MacromoleculeName: YciM / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli (E. coli)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MLELLFLLLP VAAAYGWYMG RRSAQQNKQD EANRLSRDYV AGVNFLLSNQ QDKAVDLFLD MLKEDTGTV EAHLTLGNLF RSRGEVDRAI RIHQTLMESA SLTYEQRLLA IQQLGRDYMA A GLYDRAED MFNQLTDETD FRIGALQQLL QIYQATSEWQ KAIDVAERLV ...String:
MLELLFLLLP VAAAYGWYMG RRSAQQNKQD EANRLSRDYV AGVNFLLSNQ QDKAVDLFLD MLKEDTGTV EAHLTLGNLF RSRGEVDRAI RIHQTLMESA SLTYEQRLLA IQQLGRDYMA A GLYDRAED MFNQLTDETD FRIGALQQLL QIYQATSEWQ KAIDVAERLV KLGKDKQRVE IA HFYCELA LQHMASDDLD RAMTLLKKGA AADKNSARVS IMMGRVFMAK GEYAKAVESL QRV ISQDRE LVSETLEMLQ TCYQQLGKTA EWAEFLQRAV EENTGADAEL MLADIIEARD GSEA AQVYI TRQLQRHPTM RVFHKLMDYH LNEAEEGRAK ESLMVLRDMV GEKVRSKPRY RCQKC GFTA YTLYWHCPSC RAWSTIKPIR GLDGLEHHHH HH

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Macromolecule #2: YejM

MacromoleculeName: YejM / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli (E. coli)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MVTHRQRYRE KVSQMVSWGH WFALFNILLS LVIGSRYLFI ADWPTTLAGR IYSYVSIIGH FSFLVFATY LLILFPLTFI VGSQRLMRFL SVILATAGMT LLLIDSEVFT RFHLHLNPIV W QLVINPDE NEMARDWQLM FISVPVILLL ELVFATWSWQ KLRSLTRRRR ...String:
MVTHRQRYRE KVSQMVSWGH WFALFNILLS LVIGSRYLFI ADWPTTLAGR IYSYVSIIGH FSFLVFATY LLILFPLTFI VGSQRLMRFL SVILATAGMT LLLIDSEVFT RFHLHLNPIV W QLVINPDE NEMARDWQLM FISVPVILLL ELVFATWSWQ KLRSLTRRRR FARPLAAFLF IA FIASHVV YIWADANFYR PITMQRANLP LSYPMTARRF LEKHGLLDAQ EYQRRLIEQG NPD AVSVQY PLSELRYRDM GTGQNVLLIT VDGLNYSRFE KQMPALAGFA EQNISFTRHM SSGN TTDNG IFGLFYGISP SYMDGILSTR TPAALITALN QQGYQLGLFS SDGFTSPLYR QALLS DFSM PSVRTQSDEQ TATQWINWLG RYAQEDNRWF SWVSFNGTNI DDSNQQAFAR KYSRAA GNV DDQINRVLNA LRDSGKLDNT VVIITAGRGI PLSEEEETFD WSHGHLQVPL VIHWPGT PA QRINALTDHT DLMTTLMQRL LHVSTPASEY SQGQDLFNPQ RRHYWVTAAD NDTLAITT P KKTLVLNNNG KYRTYNLRGE RVKDEKPQLS LLLQVLTDEK RFIAN

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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

Concentration6 mg/mL
BufferpH: 7.8
GridModel: C-flat-1.2/1.3 / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Average exposure time: 8.84 sec. / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 64000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: cryoSPARC (ver. 3.2)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 4.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.2) / Number images used: 175387
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.2)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.2)
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL

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