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- EMDB-42290: Cryo-EM Structure of the Helicobacter pylori CagYdAP OMC -

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Basic information

Entry
Database: EMDB / ID: EMD-42290
TitleCryo-EM Structure of the Helicobacter pylori CagYdAP OMC
Map data
Sample
  • Complex: Helicobacter pylori CagYdAP OMC
    • Protein or peptide: Type IV secretion system apparatus protein Cag3Secretion
    • Protein or peptide: Type IV secretion system apparatus protein CagMSecretion
    • Protein or peptide: Type IV secretion system apparatus protein CagTSecretion
    • Protein or peptide: Type IV secretion system apparatus protein CagXSecretion
    • Protein or peptide: Cag pathogenicity island protein (Cag7)
KeywordsT4SS / protein translocation / MEMBRANE PROTEIN
Biological speciesHelicobacter pylori 26695 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsRoberts JR
Funding support United States, 3 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)AI118932 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)AI039657 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA116087 United States
CitationJournal: Life Sci Alliance / Year: 2024
Title: Subdomains of the Cag T4SS outer membrane core complex exhibit structural independence.
Authors: Jacquelyn R Roberts / Sirena C Tran / Arwen E Frick-Cheng / Kaeli N Bryant / Chiamaka D Okoye / W Hayes McDonald / Timothy L Cover / Melanie D Ohi /
Abstract: The Cag type IV secretion system (Cag T4SS) has an important role in the pathogenesis of gastric cancer. The Cag T4SS outer membrane core complex (OMCC) is organized into three regions: a 14-fold ...The Cag type IV secretion system (Cag T4SS) has an important role in the pathogenesis of gastric cancer. The Cag T4SS outer membrane core complex (OMCC) is organized into three regions: a 14-fold symmetric outer membrane cap (OMC) composed of CagY, CagX, CagT, CagM, and Cag3; a 17-fold symmetric periplasmic ring (PR) composed of CagY and CagX; and a stalk with unknown composition. We investigated how CagT, CagM, and a conserved antenna projection (AP) region of CagY contribute to the structural organization of the OMCC. Single-particle cryo-EM analyses showed that complexes purified from Δ or Δ mutants no longer had organized OMCs, but the PRs remained structured. OMCCs purified from a CagY antenna projection mutant (CagYAP) were structurally similar to WT OMCCs, except for the absence of the α-helical antenna projection. These results indicate that CagY and CagX are sufficient for maintaining a stable PR, but the organization of the OMC requires CagY, CagX, CagM, and CagT. Our results highlight an unexpected structural independence of two major subdomains of the Cag T4SS OMCC.
History
DepositionOct 10, 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_42290.png

Downloads & links

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Map

FileDownload / File: emd_42290.map.gz / Format: CCP4 / Size: 1 GB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
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Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.08 Å/pix.
x 648 pix.
= 699.84 Å		1.08 Å/pix.
x 648 pix.
= 699.84 Å		1.08 Å/pix.
x 648 pix.
= 699.84 Å

Surface

Projections

Slices (1/3)

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Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.08 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.13
Minimum - Maximum-0.403381 - 0.7904756
Average (Standard dev.)0.0035834834 (±0.02301422)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions648648648
Spacing648648648
CellA=B=C: 699.84 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_42290_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_42290_half_map_2.map
Projections & Slices
AxesZYX

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Histogram

Histogram (log scale)

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Sample components

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Entire : Helicobacter pylori CagYdAP OMC

EntireName: Helicobacter pylori CagYdAP OMC
Components
  • Complex: Helicobacter pylori CagYdAP OMC
    • Protein or peptide: Type IV secretion system apparatus protein Cag3Secretion
    • Protein or peptide: Type IV secretion system apparatus protein CagMSecretion
    • Protein or peptide: Type IV secretion system apparatus protein CagTSecretion
    • Protein or peptide: Type IV secretion system apparatus protein CagXSecretion
    • Protein or peptide: Cag pathogenicity island protein (Cag7)

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Supramolecule #1: Helicobacter pylori CagYdAP OMC

SupramoleculeName: Helicobacter pylori CagYdAP OMC / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Helicobacter pylori 26695 (bacteria) / Strain: 26695 / Location in cell: membrane

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Macromolecule #1: Type IV secretion system apparatus protein Cag3

MacromoleculeName: Type IV secretion system apparatus protein Cag3 / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Helicobacter pylori 26695 (bacteria) / Strain: 26695
SequenceString: MFRKLATAVS LIGLLTSNTL YAKEISEADK VIKATKETKE TKKEAKRLKK EAKQRQQIPD HKKPQYVSV DDTKTQALFD IYDTLNVNDK SFGDWFGNSA LKDKTYLYAM DLLDYNNYLS I ENPIIKTR AMGTYADLII ITGSLEQVNG YYNILKALNK RNAKFVLKIN ...String:
MFRKLATAVS LIGLLTSNTL YAKEISEADK VIKATKETKE TKKEAKRLKK EAKQRQQIPD HKKPQYVSV DDTKTQALFD IYDTLNVNDK SFGDWFGNSA LKDKTYLYAM DLLDYNNYLS I ENPIIKTR AMGTYADLII ITGSLEQVNG YYNILKALNK RNAKFVLKIN ENMPYAQATF LR VPKRSDP NAHTLDKGAS IDENKLFEQQ KKMYFNYAND VICRPDDEVC SPLRDEMVAM PTS DSVTQK PNIIAPYSLY RLKETNNANE AQPSPYATQT APENSKEKLI EELIANSQLV ANEE EREKK LLAEKEKQEA ELAKYKLKDL ENQKKLKALE AELKKKNAKK PRVVEVPVSP QTSNS DETM RVVKEKENYN GLLVDKETTI KRSYEGTLIS ENSYSKKTPL NPNDLRSLEE EIKSYY IKS NGLCYTNGIN LYVKIKNDPY KEGMLCGYES VQNLLSPLKD KLKYDKQKLQ KALLKDS K

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Macromolecule #2: Type IV secretion system apparatus protein CagM

MacromoleculeName: Type IV secretion system apparatus protein CagM / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Helicobacter pylori 26695 (bacteria) / Strain: 26695
SequenceString: MLAKIVFSSL VAFGVLSAN V EQFGSFFN EI KKEQEEV AAK EDALKA RKKL LNNTH DFLED LIFR KQKIKE LMD HRAKVLS DL ENKYKKEK E ALEKETRGK ILTAKSKAYG DLEQALKDN P LYRKLLPN PY AYVLNQE TFT KEDRER LSYY YPQVK ...String:
MLAKIVFSSL VAFGVLSAN V EQFGSFFN EI KKEQEEV AAK EDALKA RKKL LNNTH DFLED LIFR KQKIKE LMD HRAKVLS DL ENKYKKEK E ALEKETRGK ILTAKSKAYG DLEQALKDN P LYRKLLPN PY AYVLNQE TFT KEDRER LSYY YPQVK TSSIF KKTT ATTKDK AQA LLQMGVF SL DEEQNKKA S RLALSYKQA IEEYSNNVSN LLSRKELDN I DYYLQLER NK FDSKAKD IAQ KATNTL IFNS ERLAF SMAID KINE KYLRGY EAF SNLLKNV KD DVELNTLT K NFTNQKLSF AQKQKLCLLV LDSFNFDTQ S KKSILKKT NE YNIFVDS DPM MSDKTT MQKE HYKIF NFFKT VVSA YRNNVA KNN PFE

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Macromolecule #3: Type IV secretion system apparatus protein CagT

MacromoleculeName: Type IV secretion system apparatus protein CagT / type: protein_or_peptide / ID: 3 / Enantiomer: LEVO
Source (natural)Organism: Helicobacter pylori 26695 (bacteria) / Strain: 26695
SequenceString: MLAKIVFSSL VAFGVLSAN V EQFGSFFN EI KKEQEEV AAK EDALKA RKKL LNNTH DFLED LIFR KQKIKE LMD HRAKVLS DL ENKYKKEK E ALEKETRGK ILTAKSKAYG DLEQALKDN P LYRKLLPN PY AYVLNQE TFT KEDRER LSYY YPQVK ...String:
MLAKIVFSSL VAFGVLSAN V EQFGSFFN EI KKEQEEV AAK EDALKA RKKL LNNTH DFLED LIFR KQKIKE LMD HRAKVLS DL ENKYKKEK E ALEKETRGK ILTAKSKAYG DLEQALKDN P LYRKLLPN PY AYVLNQE TFT KEDRER LSYY YPQVK TSSIF KKTT ATTKDK AQA LLQMGVF SL DEEQNKKA S RLALSYKQA IEEYSNNVSN LLSRKELDN I DYYLQLER NK FDSKAKD IAQ KATNTL IFNS ERLAF SMAID KINE KYLRGY EAF SNLLKNV KD DVELNTLT K NFTNQKLSF AQKQKLCLLV LDSFNFDTQ S KKSILKKT NE YNIFVDS DPM MSDKTT MQKE HYKIF NFFKT VVSA YRNNVA KNN PFE

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Macromolecule #4: Type IV secretion system apparatus protein CagX

MacromoleculeName: Type IV secretion system apparatus protein CagX / type: protein_or_peptide / ID: 4 / Enantiomer: LEVO
Source (natural)Organism: Helicobacter pylori 26695 (bacteria) / Strain: 26695
SequenceString: MKLRASVLIG ATILCLILSA CSNYAKKVVK QKNHVYTPVY NELIEKYSEI PLNDKLKDTP FMVQVKLPN YKDYLLDNKQ VVLTFKLVHH SKKITLIGDA NKILQYKNYF QANGARSDID F YLQPTLNQ KGVVMIASNY NDNPNSKEKP QTFDVLQGSQ PMLGANTKNL ...String:
MKLRASVLIG ATILCLILSA CSNYAKKVVK QKNHVYTPVY NELIEKYSEI PLNDKLKDTP FMVQVKLPN YKDYLLDNKQ VVLTFKLVHH SKKITLIGDA NKILQYKNYF QANGARSDID F YLQPTLNQ KGVVMIASNY NDNPNSKEKP QTFDVLQGSQ PMLGANTKNL HGYDVSGANN KQ VINEVAR EKAQLEKINQ YYKTLLQDKE QEYTTRKNNQ REILETLSNR AGYQMRQNVI SSE IFKNGN LNMQAKEEEV REKLQEEREN EYLRNQIRSL LSGK

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Macromolecule #5: Cag pathogenicity island protein (Cag7)

MacromoleculeName: Cag pathogenicity island protein (Cag7) / type: protein_or_peptide / ID: 5 / Enantiomer: LEVO
Source (natural)Organism: Helicobacter pylori 26695 (bacteria) / Strain: 26695
SequenceString: MGQAFFKKIV GCFCLGYLFL SSAIEAAALD IKNFNRGRVK VVNKKIAYLG DEKPITIWTS LDNVTVIQL EKDETISYIT TGFNKGWSIV PNSNHIFIQP KSVKSNLMFE KEAVNFALMT R DYQEFLKT KKLIVDAPDP KELEEQKKAL EKEKEAKEQA QKAQKDKREK ...String:
MGQAFFKKIV GCFCLGYLFL SSAIEAAALD IKNFNRGRVK VVNKKIAYLG DEKPITIWTS LDNVTVIQL EKDETISYIT TGFNKGWSIV PNSNHIFIQP KSVKSNLMFE KEAVNFALMT R DYQEFLKT KKLIVDAPDP KELEEQKKAL EKEKEAKEQA QKAQKDKREK RKEERAKNRA NL ENLTNAM SNPQNLSNNK NLSEFIKQQR ENELDQMERL EDMQEQAQAN ALKQIEELNK KQA EETIKQ RAKDKINIKT DKPQKSPEDN SIELSPSDSA WRTNLVVRTN KALYQFILRI AQKD NFASA YLTVKLEYPQ RHEVSSVIEE ELKKREEAKR QKELIKQENL NTTAYINRVM MASNE QIIN KEKIREEKQK IILDQAKALE TQYVHNALKR NPVPRNYNYY QAPEKRSKHI MPSEIF DDG TFTYFGFKNI TLQPAIFVVQ PDGKLSMTDA AIDPNMTNSG LRWYRVNEIA EKFKLIK DK ALVTVINKGY GKNPLTKNYN IKNYGELERV IKKLPLVRDK

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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

Concentration0.4 mg/mL
BufferpH: 7
VitrificationCryogen name: NITROGEN

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Electron microscopy

MicroscopeTFS 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: 1.0 µm
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 60.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2.1)
Final reconstructionApplied symmetry - Point group: C14 (14 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.2.1) / Number images used: 22463
FSC plot (resolution estimation)

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