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- EMDB-41772: S. thermodepolymerans KpsMT-KpsE with bound glycolipid - state 2 ... -

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Open data


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Basic information

Entry
Database: EMDB / ID: EMD-41772
TitleS. thermodepolymerans KpsMT-KpsE with bound glycolipid - state 2 - KpsT focused map
Map data
Sample
  • Complex: ABC transporter KpsMT in complex with polysaccharide co-polymerase KpsE bound to the glycolipid in state 2
KeywordsABC transporter / Capsular polysaccharide / MEMBRANE PROTEIN
Biological speciesCaldimonas thermodepolymerans (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsKuklewicz J / Zimmer J
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM144130 United States
CitationJournal: Nature / Year: 2024
Title: Molecular insights into capsular polysaccharide secretion.
Authors: Jeremi Kuklewicz / Jochen Zimmer /
Abstract: Capsular polysaccharides (CPSs) fortify the cell boundaries of many commensal and pathogenic bacteria. Through the ABC-transporter-dependent biosynthesis pathway, CPSs are synthesized intracellularly ...Capsular polysaccharides (CPSs) fortify the cell boundaries of many commensal and pathogenic bacteria. Through the ABC-transporter-dependent biosynthesis pathway, CPSs are synthesized intracellularly on a lipid anchor and secreted across the cell envelope by the KpsMT ABC transporter associated with the KpsE and KpsD subunits. Here we use structural and functional studies to uncover crucial steps of CPS secretion in Gram-negative bacteria. We show that KpsMT has broad substrate specificity and is sufficient for the translocation of CPSs across the inner bacterial membrane, and we determine the cell surface organization and localization of CPSs using super-resolution fluorescence microscopy. Cryo-electron microscopy analyses of the KpsMT-KpsE complex in six different states reveal a KpsE-encaged ABC transporter, rigid-body conformational rearrangements of KpsMT during ATP hydrolysis and recognition of a glycolipid inside a membrane-exposed electropositive canyon. In vivo CPS secretion assays underscore the functional importance of canyon-lining basic residues. Combined, our analyses suggest a molecular model of CPS secretion by ABC transporters.
History
DepositionAug 28, 2023-
Header (metadata) releaseApr 17, 2024-
Map releaseApr 17, 2024-
UpdateMay 8, 2024-
Current statusMay 8, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_41772.png

Downloads & links

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Map

FileDownload / File: emd_41772.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

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AxesZ (Sec.)Y (Row.)X (Col.)
1.08 Å/pix.
x 360 pix.
= 388.8 Å		1.08 Å/pix.
x 360 pix.
= 388.8 Å		1.08 Å/pix.
x 360 pix.
= 388.8 Å

Surface

Projections

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.08 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.12
Minimum - Maximum-0.98357034 - 1.6443801
Average (Standard dev.)-0.0003636915 (±0.036342014)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 388.80002 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_41772_half_map_1.map
Projections & Slices
AxesZYX

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Half map: #2

Fileemd_41772_half_map_2.map
Projections & Slices
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Histogram (log scale)

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Sample components

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Entire : ABC transporter KpsMT in complex with polysaccharide co-polymeras...

EntireName: ABC transporter KpsMT in complex with polysaccharide co-polymerase KpsE bound to the glycolipid in state 2
Components
  • Complex: ABC transporter KpsMT in complex with polysaccharide co-polymerase KpsE bound to the glycolipid in state 2

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Supramolecule #1: ABC transporter KpsMT in complex with polysaccharide co-polymeras...

SupramoleculeName: ABC transporter KpsMT in complex with polysaccharide co-polymerase KpsE bound to the glycolipid in state 2
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #2-#3, #1
Source (natural)Organism: Caldimonas thermodepolymerans (bacteria)

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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

BufferpH: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / 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: 1.8 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL / Details: AlphaFold 2
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 78036
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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