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- EMDB-42393: Cryo-EM Structure of the Helicobacter pylori dcagM PR -

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Basic information

Entry
Database: EMDB / ID: EMD-42393
TitleCryo-EM Structure of the Helicobacter pylori dcagM PR
Map data
Sample
  • Complex: Helicobacter pylori dcagM PR
    • Protein or peptide: Cag pathogenicity island protein (Cag8)
    • Protein or peptide: Cag pathogenicity island protein (Cag7)
KeywordsT4SS / protein translocation / MEMBRANE PROTEIN
Biological speciesHelicobacter pylori 26695 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 8.5 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 18, 2023-
Header (metadata) releaseApr 17, 2024-
Map releaseApr 17, 2024-
UpdateMay 1, 2024-
Current statusMay 1, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_42393.png

Downloads & links

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Map

FileDownload / File: emd_42393.map.gz / Format: CCP4 / Size: 274.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.08 Å/pix.
x 416 pix.
= 449.28 Å		1.08 Å/pix.
x 416 pix.
= 449.28 Å		1.08 Å/pix.
x 416 pix.
= 449.28 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: 0.125
Minimum - Maximum-0.29558066 - 0.5777632
Average (Standard dev.)0.0053460435 (±0.02245883)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions416416416
Spacing416416416
CellA=B=C: 449.28003 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_42393_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_42393_half_map_2.map
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Helicobacter pylori dcagM PR

EntireName: Helicobacter pylori dcagM PR
Components
  • Complex: Helicobacter pylori dcagM PR
    • Protein or peptide: Cag pathogenicity island protein (Cag8)
    • Protein or peptide: Cag pathogenicity island protein (Cag7)

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Supramolecule #1: Helicobacter pylori dcagM PR

SupramoleculeName: Helicobacter pylori dcagM PR / 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: Cag pathogenicity island protein (Cag8)

MacromoleculeName: Cag pathogenicity island protein (Cag8) / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Helicobacter pylori 26695 (bacteria)
SequenceString: MGQAFFKKIV GCFCLGYLF L SSAIEAAA LD IKNFNRG RVK VVNKKI AYLG DEKPI TIWTS LDNV TVIQLE KDE TISYITT GF NKGWSIVP N SNHIFIQPK SVKSNLMFEK EAVNFALMT R DYQEFLKT KK LIVDAPD PKE LEEQKK ALEK EKEAK ...String:
MGQAFFKKIV GCFCLGYLF L SSAIEAAA LD IKNFNRG RVK VVNKKI AYLG DEKPI TIWTS LDNV TVIQLE KDE TISYITT GF NKGWSIVP N SNHIFIQPK SVKSNLMFEK EAVNFALMT R DYQEFLKT KK LIVDAPD PKE LEEQKK ALEK EKEAK EQAQK AQKD KREKRK EER AKNRANL EN LTNAMSNP Q NLSNNKNLS EFIKQQRENE LDQMERLED M QEQAQANA LK QIEELNK KQA EETIKQ RAKD KINIK TDKPQ KSPE DNSIEL SPS DSAWRTN LV VRTNKALY Q FILRIAQKD NFASAYLTVK LEYPQRHEV S SVIEELKK RE EAKRQKE LIK QENLNT TAYI NRVMM ASNEQ IINK EKIREE KQK IILDQAK AL ETQYVHNA L KRNPVPRNY NYYQAPEKRS KHIMPSEIF D DGTFTYFG FK NITLQPA IFV VQPDGK LSMT DAAID PNMTN SGLR WYRVNE IAE KFKLIKD KA LVTVINKG Y GKNPLTKNY NIKNYGELER VIKKLPEVR D K

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Macromolecule #2: Cag pathogenicity island protein (Cag7)

MacromoleculeName: Cag pathogenicity island protein (Cag7) / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Helicobacter pylori 26695 (bacteria)
SequenceString: MNEENDKLET SKKAQQDSP Q DLSNEEAT EA NHFENLL KES KESSDH HLDN PTETQ THFDG DKSE ETQTQM DSE GNETSES SN GSLADKLF K KARKLVDNK KPFTQQKNLD EETQELNEE D DQENNEYQ EE TQTDLID DET SKKTQQ HSPQ DLSNE ...String:
MNEENDKLET SKKAQQDSP Q DLSNEEAT EA NHFENLL KES KESSDH HLDN PTETQ THFDG DKSE ETQTQM DSE GNETSES SN GSLADKLF K KARKLVDNK KPFTQQKNLD EETQELNEE D DQENNEYQ EE TQTDLID DET SKKTQQ HSPQ DLSNE EATEA NHFE NLLKES KES SDHHLDN PT ETQTNFDG D KSEETQTQM DSEGNETSES SNGSLADKL F KKARKLVD NK KPFTQQK NLD EETQEL NEED DQENN EYQEE TQTD LIDDET SKK TQQHSPQ DL SNEEATEA N HFENLLKES KESSDHHLDN PTETQTNFD G DKSEEITD DS NDQEIIK GSK KKYIIG GIVV AVLIV IILFS RSIF HYFMPL EDK SSRFSKD RN LYVNDEIQ I RQEYNRLLK ERNEKGNMID KNLFFNDDP N RTLYNYLN IA EIEDKNP LRA FYECIS NGGN YEECL KLIKD KKLQ DQMKKT LEA YNDCIKN AK TEEERIKC L DLIKDENLK KSLLNQQKVQ VALDCLKNA K TDEERNEC LK LINDPEI REK FRKELE LQKE LQEYK DCIKN AKTE AEKNKC LKG LSKEAIE RL KQQALDCL K NAKTDEERN ECLKNIPQDL QKELLADMS V KAYKDCVS KA RNEKEKQ ECE KLLTPE ARKK LEQQV LDCLK NAKT DEERKK CLK DLPKDLQ SD ILAKESLK A YKDCVSQAK TEAEKKECEK LLTPEAKKL L EEEAKESV KA YLDCVSQ AKT EAEKKE CEKL LTPEA KKKLE EAKK SVKAYL DCV SRARNEK EK KECEKLLT P EAKKLLEQQ ALDCLKNAKT DKERKKCLK D LPKDLQKK VL AKESVKA YLD CVSQAK TEAE KKECE KLLTP EARK LLEEAK KSV KAYLDCV SQ AKTEAEKK E CEKLLTPEA RKLLEEXAKE SVKAYLDCV S QAKNEAEK KE CEKLLTL ESK KKLEEA KKSV KAYLD CVSQA KTEA EKKECE KLL TPEAKKL LE QQALDCLK N AKTEADKKR CVKDLPKDLQ KKVLAKESL K AYKDCVSK AR NEKEKKE CEK LLTPEA KKLL EEAKK SVKAY LDCV SQAKTE AEK KECEKLL TP EARKLLEE A KESVKAYKD CVSKARNEKE KKECEKLLT P EAKKLLEQ QV LDCLKNA KTE ADKKRC VKDL PKDLQ KKVLA KESV KAYLDC VSR ARNEKEK KE CEKLLTPE A KKLLEEAKE SLKAYKDCLS QARNEEERR A CEKLLTPE AR KLLEQEV KKS IKAYLD CVSR ARNEK EKKEC EKLL TPEARK FLA KQVLNCL EK AGNEEERK A CLKNLPKDL QENILAKESL KAYKDCLSQ A RNEEERRA CE KLLTPEA RKL LEQEVK KSVK AYLDC VSRAR NEKE KKECEK LLT PEARKFL AK ELQQKDKA I KDCLKNADP NDRAAIMKCL DGLSDEEKL K YLQEAREK AV ADCLAMA KTD EEKRKC QNLY SDLIQ EIQNK RTQN KQNQLS KTE RLHQASE CL DNLDDPTD Q EAIEQCLEG LSDSERALIL GIKRQADEV D LIYSDLRN RK TFDNMAA KGY PLLPMD FKNG GDIAT INATN VDAD KIASDN PIY ASIEPDI AK QYETEKTI K DKNLEAKLA KALGGNKKDD DKEKSKKST A EAKAENNK ID KDVAETA KNI SEIALK NKKE KSGEF VDENG NPID DKKKAE KQD ETSPVKQ AF IGKSDPTF V LAQYTPIEI TLTSKVDATL TGIVSGVVA K DVWNMNGT MI LLDKGTK VYG NYQSVK GGTP IMTRL MIVFT KAIT PDGVII PLA NAQAAGM LG EAGVDGYV N NHFMKRIGF AVIASVVNSF LQTAPIIAL D KLIGLGKG RS ERTPEFN YAL GQAING SMQS SAQMS NQILG QLMN IPPSFY KNE GDSIKIL TM DDIDFSGV Y DVKITNKSV VDEIIKQSTK TLSREHEEI T TSPKGGN

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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 reconstructionResolution.type: BY AUTHOR / Resolution: 8.5 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: cryoSPARC (ver. 4.2.1) / Number images used: 3009
FSC plot (resolution estimation)

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