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- EMDB-41042: S. enterica WbaP in a styrene maleic acid liponanoparticle -

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Basic information

Entry
Database: EMDB / ID: EMD-41042
TitleS. enterica WbaP in a styrene maleic acid liponanoparticle
Map data
Sample
  • Complex: WbaP dimer embedded in a styrene maleic acid liponanoparticle
    • Protein or peptide: Undecaprenyl-phosphate galactose phosphotransferase
Keywordsphosphoglycosyl transferase / liponanopaticle / SMALP / O-antigen / Salmonella enterica / TRANSFERASE
Function / homology
Function and homology information


undecaprenyl-phosphate galactose phosphotransferase / undecaprenyl-phosphate galactose phosphotransferase activity / phosphotransferase activity, for other substituted phosphate groups / O antigen biosynthetic process / plasma membrane
Similarity search - Function
Undecaprenyl-phosphate galactose phosphotransferase, WbaP / Exopolysaccharide biosynthesis polyprenyl glycosylphosphotransferase / CoA-binding domain / Bacterial sugar transferase / Bacterial sugar transferase
Similarity search - Domain/homology
Undecaprenyl-phosphate galactose phosphotransferase
Similarity search - Component
Biological speciesSalmonella enterica subsp. enterica serovar Typhimurium (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.1 Å
AuthorsDodge GJ / Imperiali B
Funding support United States, 3 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM134576 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM131627 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM039334 United States
CitationJournal: Elife / Year: 2024
Title: Mapping the architecture of the initiating phosphoglycosyl transferase from O-antigen biosynthesis in a liponanoparticle.
Authors: Greg J Dodge / Alyssa J Anderson / Yi He / Weijing Liu / Rosa Viner / Barbara Imperiali /
Abstract: Bacterial cell surface glycoconjugates are critical for cell survival and for interactions between bacteria and their hosts. Consequently, the pathways responsible for their biosynthesis have ...Bacterial cell surface glycoconjugates are critical for cell survival and for interactions between bacteria and their hosts. Consequently, the pathways responsible for their biosynthesis have untapped potential as therapeutic targets. The localization of many glycoconjugate biosynthesis enzymes to the membrane represents a significant challenge for expressing, purifying, and characterizing these enzymes. Here, we leverage cutting-edge detergent-free methods to stabilize, purify, and structurally characterize WbaP, a phosphoglycosyl transferase (PGT) from the (LT2) O-antigen biosynthesis. From a functional perspective, these studies establish WbaP as a homodimer, reveal the structural elements responsible for dimerization, shed light on the regulatory role of a domain of unknown function embedded within WbaP, and identify conserved structural motifs between PGTs and functionally unrelated UDP-sugar dehydratases. From a technological perspective, the strategy developed here is generalizable and provides a toolkit for studying other classes of small membrane proteins embedded in liponanoparticles beyond PGTs.
History
DepositionJun 12, 2023-
Header (metadata) releaseFeb 14, 2024-
Map releaseFeb 14, 2024-
UpdateFeb 28, 2024-
Current statusFeb 28, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_41042.png

Downloads & links

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Map

FileDownload / File: emd_41042.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.3 Å
Density
Contour LevelBy AUTHOR: 0.149
Minimum - Maximum-1.5801021 - 1.9151361
Average (Standard dev.)-0.00013558328 (±0.022646317)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 332.8 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_41042_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_41042_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_41042_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram

Histogram (log scale)

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Sample components

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Entire : WbaP dimer embedded in a styrene maleic acid liponanoparticle

EntireName: WbaP dimer embedded in a styrene maleic acid liponanoparticle
Components
  • Complex: WbaP dimer embedded in a styrene maleic acid liponanoparticle
    • Protein or peptide: Undecaprenyl-phosphate galactose phosphotransferase

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Supramolecule #1: WbaP dimer embedded in a styrene maleic acid liponanoparticle

SupramoleculeName: WbaP dimer embedded in a styrene maleic acid liponanoparticle
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Salmonella enterica subsp. enterica serovar Typhimurium (bacteria)
Molecular weightTheoretical: 250 KDa

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Macromolecule #1: Undecaprenyl-phosphate galactose phosphotransferase

MacromoleculeName: Undecaprenyl-phosphate galactose phosphotransferase / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
EC number: undecaprenyl-phosphate galactose phosphotransferase
Source (natural)Organism: Salmonella enterica subsp. enterica serovar Typhimurium (bacteria)
Molecular weightTheoretical: 61.382699 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: GGSGSGGWSH PQFEKGGGSG GGSGGSAWSH PQFEKSSGVD LGTENLYFQS NAMDNIDNKY NPQLCKIFLA ISDLIFFNLA LWFSLGCVY FIFDQVQRFI PQDQLDTRVI THFILSVVCV GWFWIRLRHY TYRKPFWYEL KEIFRTIVIF AIFDLALIAF T KWQFSRYV ...String:
GGSGSGGWSH PQFEKGGGSG GGSGGSAWSH PQFEKSSGVD LGTENLYFQS NAMDNIDNKY NPQLCKIFLA ISDLIFFNLA LWFSLGCVY FIFDQVQRFI PQDQLDTRVI THFILSVVCV GWFWIRLRHY TYRKPFWYEL KEIFRTIVIF AIFDLALIAF T KWQFSRYV WVFCWTFALI LVPFFRALTK HLLNKLGIWK KKTIILGSGQ NARGAYSALQ SEEMMGFDVI AFFDTDASDA EI NMLPVIK DTEIIWDLNR TGDVHYILAY EYTELEKTHF WLRELSKHHC RSVTVVPSFR GLPLYNTDMS FIFSHEVMLL RIQ NNLAKR SSRFLKRTFD IVCSIMILII ASPLMIYLWY KVTRDGGPAI YGHQRVGRHG KLFPCYKFRS MVMNSQEVLK ELLA NDPIA RAEWEKDFKL KNDPRITAVG RFIRKTSLDE LPQLFNVLKG DMSLVGPRPI VSDELERYCD DVDYYLMAKP GMTGL WQVS GRNDVDYDTR VYFDSWYVKN WTLWNDIAIL FKTAKVVLRR DGAY

UniProtKB: Undecaprenyl-phosphate galactose phosphotransferase

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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

Concentration2 mg/mL
BufferpH: 8
Component:
ConcentrationNameFormula
25.0 mMHEPES
150.0 mMsodium chlorideNaClSodium chloride
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300
VitrificationCryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.75 µm / Nominal magnification: 105000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number real images: 4841 / Average electron dose: 50.13 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1266538
Startup modelType of model: NONE
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 4.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 196663
FSC plot (resolution estimation)

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