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- EMDB-10350: Type IV Coupling Complex (T4CC) from L. pneumophila. -

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Basic information

Entry
Database: EMDB / ID: EMD-10350
TitleType IV Coupling Complex (T4CC) from L. pneumophila.
Map dataCryoEM-map of T4CC hetero-pentameric core (DotLMNYZ) purified from L.pneumophila.
Sample
  • Complex: Coupling complex of the type IV secretion system
    • Protein or peptide: IcmO (DotL)
    • Protein or peptide: IcmP (DotM)
    • Protein or peptide: IcmJ (DotN)
    • Protein or peptide: DotZ
    • Protein or peptide: DotY
  • Ligand: ZINC ION
KeywordsProtein Complex / Secretion / Secretion systems / Gram-negative bacteria / type 4 secretion system / T4SS / coupling protein / T4CC / DotL / DotM / DotN / DotY / DotZ / MEMBRANE PROTEIN
Function / homology
Function and homology information


protein transport / metal ion binding / plasma membrane / cytoplasm
Similarity search - Function
: / : / : / TraD/TraG, TraM recognition site / TraM recognition site of TraD and TraG / : / : / HNH nucleases / HNH nuclease / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Type 4 apparatus protein DotZ / Type 4 apparatus protein DotN / Type 4 coupling protein DotL / Type 4 apparatus protein DotM / Type 4 apparatus protein DotY
Similarity search - Component
Biological speciesLegionella pneumophila (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsMace K / Meir A / Lukoyanova N / Waksman G
Funding support United Kingdom, 1 items
OrganizationGrant numberCountry
European Research Council321630 United Kingdom
CitationJournal: Nat Commun / Year: 2020
Title: Mechanism of effector capture and delivery by the type IV secretion system from Legionella pneumophila.
Authors: Amit Meir / Kevin Macé / Natalya Lukoyanova / David Chetrit / Manuela K Hospenthal / Adam Redzej / Craig Roy / Gabriel Waksman /
Abstract: Legionella pneumophila is a bacterial pathogen that utilises a Type IV secretion (T4S) system to inject effector proteins into human macrophages. Essential to the recruitment and delivery of ...Legionella pneumophila is a bacterial pathogen that utilises a Type IV secretion (T4S) system to inject effector proteins into human macrophages. Essential to the recruitment and delivery of effectors to the T4S machinery is the membrane-embedded T4 coupling complex (T4CC). Here, we purify an intact T4CC from the Legionella membrane. It contains the DotL ATPase, the DotM and DotN proteins, the chaperone module IcmSW, and two previously uncharacterised proteins, DotY and DotZ. The atomic resolution structure reveals a DotLMNYZ hetero-pentameric core from which the flexible IcmSW module protrudes. Six of these hetero-pentameric complexes may assemble into a 1.6-MDa hexameric nanomachine, forming an inner membrane channel for effectors to pass through. Analysis of multiple cryo EM maps, further modelling and mutagenesis provide working models for the mechanism for binding and delivery of two essential classes of Legionella effectors, depending on IcmSW or DotM, respectively.
History
DepositionOct 2, 2019-
Header (metadata) releaseMay 13, 2020-
Map releaseMay 13, 2020-
UpdateJul 10, 2024-
Current statusJul 10, 2024Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 7
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 7
  • Imaged by UCSF Chimera
  • Download
  • Surface view with fitted model
  • Atomic models: PDB-6sz9
  • Surface level: 7
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_10350.png

Downloads & links

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Map

FileDownload / File: emd_10350.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryoEM-map of T4CC hetero-pentameric core (DotLMNYZ) purified from L.pneumophila.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.05 Å/pix.
x 300 pix.
= 313.5 Å		1.05 Å/pix.
x 300 pix.
= 313.5 Å		1.05 Å/pix.
x 300 pix.
= 313.5 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.045 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 7.0 / Movie #1: 7
Minimum - Maximum-29.188368000000001 - 42.54036
Average (Standard dev.)-0.00016760475 (±0.841204)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 313.5 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0451.0451.045
M x/y/z300300300
origin x/y/z0.0000.0000.000
length x/y/z313.500313.500313.500
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS300300300
D min/max/mean-29.18842.540-0.000

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Supplemental data

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Half map: Half map A

Fileemd_10350_half_map_1.map
AnnotationHalf map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map B

Fileemd_10350_half_map_2.map
AnnotationHalf map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Coupling complex of the type IV secretion system

EntireName: Coupling complex of the type IV secretion system
Components
  • Complex: Coupling complex of the type IV secretion system
    • Protein or peptide: IcmO (DotL)
    • Protein or peptide: IcmP (DotM)
    • Protein or peptide: IcmJ (DotN)
    • Protein or peptide: DotZ
    • Protein or peptide: DotY
  • Ligand: ZINC ION

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Supramolecule #1: Coupling complex of the type IV secretion system

SupramoleculeName: Coupling complex of the type IV secretion system / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#5
Source (natural)Organism: Legionella pneumophila (bacteria)

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Macromolecule #1: IcmO (DotL)

MacromoleculeName: IcmO (DotL) / type: protein_or_peptide / ID: 1 / Details: C-terminal Streptavidin tag / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Legionella pneumophila (bacteria)
Molecular weightTheoretical: 87.600992 KDa
Recombinant expressionOrganism: Legionella pneumophila (bacteria)
SequenceString: MMRGIDSRHE LDPTLLLRDT RTFTQRLADF FADPTNISIV LISLAAVSYY FSEAATFLLI MGGIFFLYSY TRKQKLPFRL PQISRAKDY NDLKPGINKP NIARGITFFG NDRKTGEELW FANDDMRTHA LIFGSTGSGK TETLVSLSYN ALVQGSGFIY V DGKGDNSL ...String:
MMRGIDSRHE LDPTLLLRDT RTFTQRLADF FADPTNISIV LISLAAVSYY FSEAATFLLI MGGIFFLYSY TRKQKLPFRL PQISRAKDY NDLKPGINKP NIARGITFFG NDRKTGEELW FANDDMRTHA LIFGSTGSGK TETLVSLSYN ALVQGSGFIY V DGKGDNSL YAKVFSMVRS MGREDDLLLI NFMTGARDIV GPQEKRLSNT LNPFCQGSSS MLTQLVVSLM GSSGQSSDGD MW KGRAIAF VEALMRLLVY MRDEGAILLD ANTIRNYFDL QRLESIVIDK VFPRDDQESV NIETIPKLVT DPLRNYLNTL PGY NKEKKG KQVSQVLEQH GFITMQLVRS FSSLADTYGH IIRTNLAEVD FKDVVLNRRI LVVLLPALEK SPDELSNLGK IIVS SLKAM MAAGLGEEVE GDYRDVILRK PTNAPTPYMC ILDEYGYYAV QGFAVVPAQA RSLGFSAIFA GQDLPAFQKA SKEEA ASIG ANTNIKICMK LEDPTETWDF FTKTAGEAYV TKVDSFQTKE TSIANSYMDT KSSSFEKRAR VDLLDLKEQT EGEAHI FFK SKIVRARMFY ANPKPVKQLK INQFLKVEPP PDDYLMKLQK QLASFQSILE SGDLSINKAV ENEEITLISK ALKESTI VE PIERGVAALI AFHGQNEPEP VEDIVEEEVE GALTIFSKLR IDPNAPPILV ADKEVFSEPL LPINETRNQM ITIERLAG A KDKYAGTVAN ELIKDFQIAT SYPPEERDVI DVQELTGIIR DLSAKISAER EKANKKAAEE LT

UniProtKB: Type 4 coupling protein DotL

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Macromolecule #2: IcmP (DotM)

MacromoleculeName: IcmP (DotM) / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Legionella pneumophila (bacteria)
Molecular weightTheoretical: 43.858879 KDa
Recombinant expressionOrganism: Legionella pneumophila (bacteria)
SequenceString: MYIEMAQQQQ QSGSDNSMAP VWIVILLFIT AYFVWALAHQ YIVSFVFTIN IWQARLVNLF LNNQLLANQI YLMQTLDPNT VNWDQMVTV MRAVGDYMRY PVICILVVLA FVLYNSNVTL KYRKTYDMKS LRAQEQFNWP AIMPIVKEDL VSQDVNKGPW A MALTPMEF ...String:
MYIEMAQQQQ QSGSDNSMAP VWIVILLFIT AYFVWALAHQ YIVSFVFTIN IWQARLVNLF LNNQLLANQI YLMQTLDPNT VNWDQMVTV MRAVGDYMRY PVICILVVLA FVLYNSNVTL KYRKTYDMKS LRAQEQFNWP AIMPIVKEDL VSQDVNKGPW A MALTPMEF ARKYNLLRKD DALLDNPVPG EEMTAGIRRG DAKRVFTMQL GPYWDGFERC SPQAYALSAV FMARMNRDRD AA NNILKVL DKTFVDGKPD FSVARPVMKK YQNSELVQEV VAKHAYVLTV IASLLEAARE DGVVPSSEFL WLKPVDRRLW YML NCVGRQ TPYSEVAGPF AHWKAEKEMG RRSLVPMIDE AIRALEIAVK EVRLTPRQME ELEP

UniProtKB: Type 4 apparatus protein DotM

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Macromolecule #3: IcmJ (DotN)

MacromoleculeName: IcmJ (DotN) / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Legionella pneumophila (bacteria)
Molecular weightTheoretical: 23.762971 KDa
Recombinant expressionOrganism: Legionella pneumophila (bacteria)
SequenceString: MADNQQRCEL KLIASPGSWR LYSARKIDER FKSYEQKIFQ RDRYTCQFCG FQARLYQDIV NLDGDYTNNR LSNLVTACCF CAQCFFVES VGVGGYGGGT LIYLPELTQA ELNSLCHVLF CAITNDTGYK SSAQNIYRSF KFRSQIVEEK FGEGTSDPAI F GQLMIDSG ...String:
MADNQQRCEL KLIASPGSWR LYSARKIDER FKSYEQKIFQ RDRYTCQFCG FQARLYQDIV NLDGDYTNNR LSNLVTACCF CAQCFFVES VGVGGYGGGT LIYLPELTQA ELNSLCHVLF CAITNDTGYK SSAQNIYRSF KFRSQIVEEK FGEGTSDPAI F GQLMIDSG VNSEEIREKL FKNIRLLPSR AKFRKQIEKW AASALEEIAD

UniProtKB: Type 4 apparatus protein DotN

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Macromolecule #4: DotZ

MacromoleculeName: DotZ / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Legionella pneumophila (bacteria)
Molecular weightTheoretical: 33.887512 KDa
Recombinant expressionOrganism: Legionella pneumophila (bacteria)
SequenceString: MDEIKKDDEL SQWLSTYGTI TAERILGRYN ISLPQDEILE AINIPSSFYR HLLQIPLKNV LNGIVIQQAS DYHVYAQKLL IDYLLSGES SKEPDSQGAG TRESLEDERQ RLVQLGDEFH KLELEQDNLI ASSQASLMKI SIDWNTKLET TLSKLNSLYK N TNSKIKKN ...String:
MDEIKKDDEL SQWLSTYGTI TAERILGRYN ISLPQDEILE AINIPSSFYR HLLQIPLKNV LNGIVIQQAS DYHVYAQKLL IDYLLSGES SKEPDSQGAG TRESLEDERQ RLVQLGDEFH KLELEQDNLI ASSQASLMKI SIDWNTKLET TLSKLNSLYK N TNSKIKKN AIRKALIKAF IHCDLVKDQS QKNKYQLIDK LNQTLAVSVG AELKESILTN LSELFQILEA LNTKLDEFTD RT NHLSQQA KSFRTQFYEV ILRIIELIKL LPEYKIDPAQ DAINREPLYF DRTIGER

UniProtKB: Type 4 apparatus protein DotZ

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Macromolecule #5: DotY

MacromoleculeName: DotY / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Legionella pneumophila (bacteria)
Molecular weightTheoretical: 25.550957 KDa
Recombinant expressionOrganism: Legionella pneumophila (bacteria)
SequenceString: MPKYTLPTRD ALLKAMQVGE TSIEAAEYMA TRFEQILTKA KLLPECNDML EKIKEYAQFV KFKLLSSAQV WSGQERPTSD YQNTQENKA EFLASHLEGL PSGLKLEVAI GDDAKILRGF SSNGKMVEGD QLKTMDGLLE GWLAKNSLAI SGGAVVKIDN T GNQTKVDP ...String:
MPKYTLPTRD ALLKAMQVGE TSIEAAEYMA TRFEQILTKA KLLPECNDML EKIKEYAQFV KFKLLSSAQV WSGQERPTSD YQNTQENKA EFLASHLEGL PSGLKLEVAI GDDAKILRGF SSNGKMVEGD QLKTMDGLLE GWLAKNSLAI SGGAVVKIDN T GNQTKVDP QEIRQLINDS EKGVAKYFAD KGVGMEVAQR TYQEPKALET KREEIRQEIE SGAEAPTTQS IR

UniProtKB: Type 4 apparatus protein DotY

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Macromolecule #6: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 6 / Number of copies: 1 / Formula: ZN
Molecular weightTheoretical: 65.409 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

Concentration0.175 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
0.2 MNaClsodium chloride
50.0 mMTris-HCltris aminomethane hydrochloride
20.0 mMMgSO4Magnesium sulfate
1.0 mMEDTAEthylenediaminetetraacetic acid
GridModel: UltrAuFoil / Material: GOLD / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 94 % / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 54.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: OTHER
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL / In silico model: CryoSPARC ab-initio
Final reconstructionNumber classes used: 2 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 2) / Number images used: 219593
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3)
Final angle assignmentType: OTHER
Details: Stochastic gradient descent (SGD) and branch-and-bound maximum likelihood optimization algorithms.
Final 3D classificationNumber classes: 4 / Software - Name: RELION (ver. 3)
FSC plot (resolution estimation)

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