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- EMDB-15701: Outer membrane secretin pore of the type 3 secretion system of Sh... -

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Basic information

Entry
Database: EMDB / ID: EMD-15701
TitleOuter membrane secretin pore of the type 3 secretion system of Shigella flexneri
Map dataMain map sharpened and masked
Sample
  • Complex: Needle complex of the type 3 secretion system
    • Complex: Outer membrane secretin pore of the type 3 secretion system
      • Protein or peptide: Outer membrane protein MxiD
Keywordstype 3 secretion / secretin / outer membrane ring / shigella / infection / PROTEIN TRANSPORT
Function / homology
Function and homology information


type III protein secretion system complex / protein secretion by the type III secretion system / cell outer membrane
Similarity search - Function
: / SPI-1 type 3 secretion system secretin, N0 domain / Type III secretion system outer membrane pore YscC/HrcC / Bacterial type II secretion system protein D signature. / Type II secretion system protein GspD, conserved site / : / NolW-like / NolW-like superfamily / Bacterial type II/III secretion system short domain / Type II/III secretion system / Bacterial type II and III secretion system protein
Similarity search - Domain/homology
Type 3 secretion system secretin
Similarity search - Component
Biological speciesShigella flexneri (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.42 Å
AuthorsLunelli M
Funding support Germany, 1 items
OrganizationGrant numberCountry
Helmholtz Association Germany
CitationJournal: Protein Sci / Year: 2023
Title: Integrative structural analysis of the type III secretion system needle complex from Shigella flexneri.
Authors: Lara Flacht / Michele Lunelli / Karol Kaszuba / Zhuo Angel Chen / Francis J O' Reilly / Juri Rappsilber / Jan Kosinski / Michael Kolbe /
Abstract: The type III secretion system (T3SS) is a large, transmembrane protein machinery used by various pathogenic gram-negative bacteria to transport virulence factors into the host cell during infection. ...The type III secretion system (T3SS) is a large, transmembrane protein machinery used by various pathogenic gram-negative bacteria to transport virulence factors into the host cell during infection. Understanding the structure of T3SSs is crucial for future developments of therapeutics that could target this system. However, much of the knowledge about the structure of T3SS is available only for Salmonella, and it is unclear how this large assembly is conserved across species. Here, we combined cryo-electron microscopy, cross-linking mass spectrometry, and integrative modeling to determine the structure of the T3SS needle complex from Shigella flexneri. We show that the Shigella T3SS exhibits unique features distinguishing it from other structurally characterized T3SSs. The secretin pore complex adopts a new fold of its C-terminal S domain and the pilotin MxiM[SctG] locates around the outer surface of the pore. The export apparatus structure exhibits a conserved pseudohelical arrangement but includes the N-terminal domain of the SpaS[SctU] subunit, which was not present in any of the previously published virulence-related T3SS structures. Similar to other T3SSs, however, the apparatus is anchored within the needle complex by a network of flexible linkers that either adjust conformation to connect to equivalent patches on the secretin oligomer or bind distinct surface patches at the same height of the export apparatus. The conserved and unique features delineated by our analysis highlight the necessity to analyze T3SS in a species-specific manner, in order to fully understand the underlying molecular mechanisms of these systems. The structure of the type III secretion system from Shigella flexneri delineates conserved and unique features, which could be used for the development of broad-range therapeutics.
History
DepositionAug 31, 2022-
Header (metadata) releaseMar 1, 2023-
Map releaseMar 1, 2023-
UpdateNov 13, 2024-
Current statusNov 13, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_15701.png

Downloads & links

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Map

FileDownload / File: emd_15701.map.gz / Format: CCP4 / Size: 421.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMain map sharpened and masked
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
1.38 Å/pix.
x 480 pix.
= 664.171 Å		1.38 Å/pix.
x 480 pix.
= 664.171 Å		1.38 Å/pix.
x 480 pix.
= 664.171 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.38369 Å
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Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.07
Minimum - Maximum-0.23210317 - 0.41871837
Average (Standard dev.)0.00010633163 (±0.0044784253)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions480480480
Spacing480480480
CellA=B=C: 664.1712 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_15701_msk_1.map
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Additional map: Main map unsharpened and unmasked

Fileemd_15701_additional_1.map
AnnotationMain map unsharpened and unmasked
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Half map: #2

Fileemd_15701_half_map_1.map
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Half map: #1

Fileemd_15701_half_map_2.map
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Sample components

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Entire : Needle complex of the type 3 secretion system

EntireName: Needle complex of the type 3 secretion system
Components
  • Complex: Needle complex of the type 3 secretion system
    • Complex: Outer membrane secretin pore of the type 3 secretion system
      • Protein or peptide: Outer membrane protein MxiD

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Supramolecule #1: Needle complex of the type 3 secretion system

SupramoleculeName: Needle complex of the type 3 secretion system / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Shigella flexneri (bacteria)

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Supramolecule #2: Outer membrane secretin pore of the type 3 secretion system

SupramoleculeName: Outer membrane secretin pore of the type 3 secretion system
type: complex / ID: 2 / Parent: 1 / Macromolecule list: all
Details: Pore formed by the oligomerization of the domains N3, Secretin and S of MxiD
Source (natural)Organism: Shigella flexneri (bacteria)

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Macromolecule #1: Outer membrane protein MxiD

MacromoleculeName: Outer membrane protein MxiD / type: protein_or_peptide / ID: 1 / Number of copies: 15 / Enantiomer: LEVO
Source (natural)Organism: Shigella flexneri (bacteria)
Molecular weightTheoretical: 63.230414 KDa
SequenceString: MKKFNIKSLT LLIVLLPLIV NANNIDSHLL EQNDIAKYVA QSDTVGSFFE RFSALLNYPI VVSKQAAKKR ISGEFDLSNP EEMLEKLTL LVGLIWYKDG NALYIYDSGE LISKVILLEN ISLNYLIQYL KDANLYDHRY PIRGNISDKT FYISGPPALV E LVANTATL ...String:
MKKFNIKSLT LLIVLLPLIV NANNIDSHLL EQNDIAKYVA QSDTVGSFFE RFSALLNYPI VVSKQAAKKR ISGEFDLSNP EEMLEKLTL LVGLIWYKDG NALYIYDSGE LISKVILLEN ISLNYLIQYL KDANLYDHRY PIRGNISDKT FYISGPPALV E LVANTATL LDKQVSSIGT DKVNFGVIKL KNTFVSDRTY NMRGEDIVIP GVATVVERLL NNGKALSNRQ AQNDPMPPFN IT QKVSEDS NDFSFSSVTN SSILEDVSLI AYPETNSILV KGNDQQIQII RDIITQLDVA KRHIELSLWI IDIDKSELNN LGV NWQGTA SFGDSFGASF NMSSSASIST LDGNKFIASV MALNQKKKAN VVSRPVILTQ ENIPAIFDNN RTFYVSLVGE RNSS LEHVT YGTLINVIPR FSSRGQIEMS LTIEDGTGNS QSNYNYNNEN TSVLPEVGRT KISTIARVPQ GKSLLIGGYT HETNS NEII SIPFLSSIPV IGNVFKYKTS NISNIVRVFL IQPREIKESS YYNTAEYKSL ISEREIQKTT QIIPSETTLL EDEKSL VSY LNY

UniProtKB: Type 3 secretion system secretin

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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: 8
GridModel: Quantifoil R2/1 / Material: COPPER / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON II (4k x 4k) / Detector mode: INTEGRATING / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Digitization - Frames/image: 1-6 / Number grids imaged: 1 / Number real images: 5238 / Average exposure time: 1.5 sec. / Average electron dose: 25.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 4.0 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 101179
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

Startup modelType of model: INSILICO MODEL
In silico model: Homology model obtained from Salmonella secretin
Final reconstructionApplied symmetry - Point group: C15 (15 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.42 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0)
Details: Focused reconstruction obtained after partial signal subtraction.
Number images used: 69589
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Overall B value: 72.8
Output model

PDB-8axl:
Outer membrane secretin pore of the type 3 secretion system of Shigella flexneri

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