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- EMDB-52896: Cryo-EM structure of Shigella flexneri LptDE bound by a Bicyclic ... -

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Basic information

Entry
Database: EMDB / ID: EMD-52896
TitleCryo-EM structure of Shigella flexneri LptDE bound by a Bicyclic peptide molecule (Compound 16)
Map data
Sample
  • Complex: Cryo-EM structure of S. flexneri LptDE complex bound by a Bicyclic peptide molecule (Compound 8)
    • Protein or peptide: LPS-assembly protein LptD
    • Protein or peptide: LPS-assembly lipoprotein LptE
  • Protein or peptide: Compound 16 - Bicyclic Peptide Binder
  • Ligand: DODECYL-BETA-D-MALTOSIDE
  • Ligand: PALMITIC ACID
  • Ligand: (2S)-3-hydroxypropane-1,2-diyl dihexadecanoate
  • Ligand: 1,3,5-tris(2-chloroethylsulfonyl)-1,3,5-triazinane
KeywordsLipid transport / Outer membrane protein / MEMBRANE PROTEIN
Function / homology
Function and homology information


transporter complex / lipopolysaccharide transport / Gram-negative-bacterium-type cell outer membrane assembly / cell outer membrane / lipopolysaccharide binding
Similarity search - Function
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 / LPS-assembly protein LptD
Similarity search - Component
Biological speciesShigella flexneri (bacteria) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.54 Å
AuthorsAllyjaun S / Newman H / Chirgadze DY / Hardwick SW / Hubbard J / van den Berg B / Dunbar E
Funding support United Kingdom, 2 items
OrganizationGrant numberCountry
Innovate UK10086683 United Kingdom
Medical Research Council (MRC, United Kingdom)U-016275 IAA-CiC United Kingdom
CitationJournal: J Med Chem / Year: 2025
Title: High-Throughput Identification and Characterization of LptDE-Binding Bicycle Peptides Using Phage Display and Cryo-EM.
Authors: Shenaz Allyjaun / Emily Dunbar / Steven W Hardwick / Sarah Newell / Finn Holding / Catherine E Rowland / Megan A St Denis / Simone Pellegrino / Gustavo Arruda Bezerra / Nikolaos Bournakas / ...Authors: Shenaz Allyjaun / Emily Dunbar / Steven W Hardwick / Sarah Newell / Finn Holding / Catherine E Rowland / Megan A St Denis / Simone Pellegrino / Gustavo Arruda Bezerra / Nikolaos Bournakas / Dimitri Y Chirgadze / Lee Cooper / Giulia Paris / Nick Lewis / Peter Brown / Michael J Skynner / Michael J Dawson / Paul Beswick / Julia Hubbard / Bert van den Berg / Hector Newman /
Abstract: The lipopolysaccharide (LPS) transport (Lpt) system in Gram-negative bacteria maintains the integrity of the asymmetric bacterial outer membrane (OM). LPS biogenesis systems are essential in most ...The lipopolysaccharide (LPS) transport (Lpt) system in Gram-negative bacteria maintains the integrity of the asymmetric bacterial outer membrane (OM). LPS biogenesis systems are essential in most Gram-negative bacteria, with LptDE responsible for the delivery of LPS to the outer leaflet of the OM. As an externally accessible, essential protein, LptDE offers a promising target for inhibitor development without the need for cellular penetration. However, there are no direct inhibitors of LptDE, and drug discovery is made challenging since it is a membrane target without a conventional active site. Here, the bicycle phage display platform was used in combination with cryogenic-electron microscopy (cryo-EM) and surface plasmon resonance to identify and map bicyclic peptide binders to LptDE (SfLptDE). Four distinct epitopes with unique bicycle molecule binding motifs were identified across the SfLptD β-barrel. This method represents a streamlined workflow for the identification and prioritization of hit molecules against LptDE.
History
DepositionFeb 25, 2025-
Header (metadata) releaseOct 15, 2025-
Map releaseOct 15, 2025-
UpdateOct 15, 2025-
Current statusOct 15, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_52896.png

Downloads & links

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Map

FileDownload / File: emd_52896.map.gz / Format: CCP4 / Size: 166.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.73 Å/pix.
x 352 pix.
= 256.608 Å		0.73 Å/pix.
x 352 pix.
= 256.608 Å		0.73 Å/pix.
x 352 pix.
= 256.608 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.729 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.105
Minimum - Maximum-0.21304396 - 0.45127812
Average (Standard dev.)0.00068280357 (±0.010969199)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions352352352
Spacing352352352
CellA=B=C: 256.608 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_52896_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_52896_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 : Cryo-EM structure of S. flexneri LptDE complex bound by a Bicycli...

EntireName: Cryo-EM structure of S. flexneri LptDE complex bound by a Bicyclic peptide molecule (Compound 8)
Components
  • Complex: Cryo-EM structure of S. flexneri LptDE complex bound by a Bicyclic peptide molecule (Compound 8)
    • Protein or peptide: LPS-assembly protein LptD
    • Protein or peptide: LPS-assembly lipoprotein LptE
  • Protein or peptide: Compound 16 - Bicyclic Peptide Binder
  • Ligand: DODECYL-BETA-D-MALTOSIDE
  • Ligand: PALMITIC ACID
  • Ligand: (2S)-3-hydroxypropane-1,2-diyl dihexadecanoate
  • Ligand: 1,3,5-tris(2-chloroethylsulfonyl)-1,3,5-triazinane

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Supramolecule #1: Cryo-EM structure of S. flexneri LptDE complex bound by a Bicycli...

SupramoleculeName: Cryo-EM structure of S. flexneri LptDE complex bound by a Bicyclic peptide molecule (Compound 8)
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Shigella flexneri (bacteria)
Molecular weightTheoretical: 190 KDa

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Macromolecule #1: LPS-assembly protein LptD

MacromoleculeName: LPS-assembly protein LptD / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Shigella flexneri (bacteria)
Molecular weightTheoretical: 87.135469 KDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria)
SequenceString: ADLASQCMLG VPSYDRPLVQ GDTNDLPVTI NADHAKGDYP DDAVFTGSVD IMQGNSRLQA DEVQLHQKEA PGQPEPVRTV DALGNVHYD DNQVILKGPK GWANLNTKDT NVWEGDYQMV GRQGRGKADL MKQRGENRYT ILDNGSFTSC LPGSDTWSVV G SEIIHDRE ...String:
ADLASQCMLG VPSYDRPLVQ GDTNDLPVTI NADHAKGDYP DDAVFTGSVD IMQGNSRLQA DEVQLHQKEA PGQPEPVRTV DALGNVHYD DNQVILKGPK GWANLNTKDT NVWEGDYQMV GRQGRGKADL MKQRGENRYT ILDNGSFTSC LPGSDTWSVV G SEIIHDRE EQVAEIWNAR FKVGPVPIFY SPYLQLPVGD KRRSGFLIPN AKYTTTNYFE FYLPYYWNIA PNMDATITPH YM HRRGNIM WENEFRYLSQ AGAGLMELDY LPSDKVYEDE HPNDDSSRRW LFYWNHSGVM DQVWRFNVDY TKVSDPSYFN DFD NKYGSS TDGYATQKFS VGYAVQNFNA TVSTKQFQVF SEQNTSSYSA EPQLDVNYYQ NDVGPFDTRI YGQAVHFVNT RDDM PEATR VHLEPTINLP LSNNWGSINT EAKFLATHYQ QTNLDWYNSR NTTKLDESVN RVMPQFKVDG KMVFERDMEM LAPGY TQTL EPRAQYLYVP YRDQSDIYNY DSSLLQSDYS GLFRDRTYGG LDRIASANQV TTGVTSRIYD DAAVERFNIS VGQIYY FTE SRTGDDNITW ENDDKTGSLV WAGDTYWRIS ERWGLRGGIQ YDTRLDNVAT SNSSIEYRRD EDRLVQLNYH YASPEYI QA TLPKYYSTAE QYKNGISQVG AVASRPIADR WSIVGAYYYD TNANKQADSM LGVQYSSCCY AIRVGYERKL NGWDNDKQ H AVYDNAIGFN IELRGLSSNY GLGTQEMLRS NILPYQNTL

UniProtKB: LPS-assembly protein LptD

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Macromolecule #2: LPS-assembly lipoprotein LptE

MacromoleculeName: LPS-assembly lipoprotein LptE / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Shigella flexneri (bacteria)
Molecular weightTheoretical: 17.002602 KDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria)
SequenceString:
CGWHLRDTTQ VPSTMKVMIL DSGDPNGPLS RAVRNQLRLN GVELLDKETT RKDVPSLRLG KVSIAKDTAS VFRNGQTAEY QMIMTVNAT VLIPGRDIYP ISAKVFRSFF DNPQMALAKD NEQDMIVKEM YDRAAEQLIR KLPSIRAADI RS

UniProtKB: LPS-assembly lipoprotein LptE

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Macromolecule #3: Compound 16 - Bicyclic Peptide Binder

MacromoleculeName: Compound 16 - Bicyclic Peptide Binder / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 1.847186 KDa
SequenceString:
ACEWCIFWYD PKLCA(NH2)

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Macromolecule #4: DODECYL-BETA-D-MALTOSIDE

MacromoleculeName: DODECYL-BETA-D-MALTOSIDE / type: ligand / ID: 4 / Number of copies: 2 / Formula: LMT
Molecular weightTheoretical: 510.615 Da
Chemical component information

ChemComp-LMT:
DODECYL-BETA-D-MALTOSIDE / detergent*YM

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Macromolecule #5: PALMITIC ACID

MacromoleculeName: PALMITIC ACID / type: ligand / ID: 5 / Number of copies: 1 / Formula: PLM
Molecular weightTheoretical: 256.424 Da
Chemical component information

ChemComp-PLM:
PALMITIC ACID

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Macromolecule #6: (2S)-3-hydroxypropane-1,2-diyl dihexadecanoate

MacromoleculeName: (2S)-3-hydroxypropane-1,2-diyl dihexadecanoate / type: ligand / ID: 6 / Number of copies: 1 / Formula: Z41
Molecular weightTheoretical: 568.911 Da
Chemical component information

ChemComp-Z41:
(2S)-3-hydroxypropane-1,2-diyl dihexadecanoate

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Macromolecule #7: 1,3,5-tris(2-chloroethylsulfonyl)-1,3,5-triazinane

MacromoleculeName: 1,3,5-tris(2-chloroethylsulfonyl)-1,3,5-triazinane / type: ligand / ID: 7 / Number of copies: 1 / Formula: A1I4O
Molecular weightTheoretical: 466.81 Da

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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

Concentration8 mg/mL
BufferpH: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: TFS FALCON 4i (4k x 4k) / Average electron dose: 12.06 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.8 µm / Nominal defocus min: 0.6 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.54 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 168619
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. v4.6.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. v4.6.0)
FSC plot (resolution estimation)

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