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- PDB-6x6k: Cryo-EM Structure of the Helicobacter pylori dCag3 OMC -

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Basic information

Entry
Database: PDB / ID: 6x6k
TitleCryo-EM Structure of the Helicobacter pylori dCag3 OMC
Components
  • Cag pathogenicity island protein
  • Cag pathogenicity island protein (Cag7)
  • Type IV secretion system apparatus protein CagXSecretion
KeywordsPROTEIN TRANSPORT / Helicobacter / T4SS / Secretion
Function / homology
Function and homology information


Cag pathogenicity island protein Cag12 / Cag pathogenicity island protein Cag12 / CagY type 1 repeat / CagY type 1 repeat / DC-EC / DC-EC Repeat / Type IV secretion system CagX conjugation protein / Conjugal transfer, TrbG/VirB9/CagX / VirB9/CagX/TrbG, C-terminal / VirB9/CagX/TrbG, C-terminal domain superfamily ...Cag pathogenicity island protein Cag12 / Cag pathogenicity island protein Cag12 / CagY type 1 repeat / CagY type 1 repeat / DC-EC / DC-EC Repeat / Type IV secretion system CagX conjugation protein / Conjugal transfer, TrbG/VirB9/CagX / VirB9/CagX/TrbG, C-terminal / VirB9/CagX/TrbG, C-terminal domain superfamily / Conjugal transfer protein / Type IV secretion system, VirB10/TrbI / Bacterial conjugation TrbI-like protein / Type IV secretion system, VirB10 / TraB / TrbI / Prokaryotic membrane lipoprotein lipid attachment site profile.
Similarity search - Domain/homology
Type IV secretion system apparatus protein CagX / Cag pathogenicity island protein (Cag7) / Cag pathogenicity island protein
Similarity search - Component
Biological speciesHelicobacter pylori (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsSheedlo, M.J. / Chung, J.M. / Sawhney, N. / Durie, C.L. / Cover, T.L. / Ohi, M.D. / Lacy, D.B.
Funding support United States, 7items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)AI118932 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA116087 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)AI039657 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM103310 United States
National Institutes of Health/National Institute of Diabetes and Digestive and Kidney Disease (NIH/NIDDK)2T32DK007673 United States
National Institutes of Health/Office of the DirectorS10OD020011 United States
Other government1I01BX004447 United States
CitationJournal: Elife / Year: 2020
Title: Cryo-EM reveals species-specific components within the Cag type IV secretion system core complex.
Authors: Michael J Sheedlo / Jeong Min Chung / Neha Sawhney / Clarissa L Durie / Timothy L Cover / Melanie D Ohi / D Borden Lacy /
Abstract: The pathogenesis of -associated gastric cancer is dependent on delivery of CagA into host cells through a type IV secretion system (T4SS). The Cag T4SS includes a large membrane-spanning core ...The pathogenesis of -associated gastric cancer is dependent on delivery of CagA into host cells through a type IV secretion system (T4SS). The Cag T4SS includes a large membrane-spanning core complex containing five proteins, organized into an outer membrane cap (OMC), a periplasmic ring (PR) and a stalk. Here, we report cryo-EM reconstructions of a core complex lacking Cag3 and an improved map of the wild-type complex. We define the structures of two unique species-specific components (Cag3 and CagM) and show that Cag3 is structurally similar to CagT. Unexpectedly, components of the OMC are organized in a 1:1:2:2:5 molar ratio (CagY:CagX:CagT:CagM:Cag3). CagX and CagY are components of both the OMC and the PR and bridge the symmetry mismatch between these regions. These results reveal that assembly of the T4SS core complex is dependent on incorporation of interwoven species-specific components.
History
DepositionMay 28, 2020Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 7, 2020Provider: repository / Type: Initial release
Revision 1.1Apr 14, 2021Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID
Revision 1.2Mar 6, 2024Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / database_2
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

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Assembly

Deposited unit
AT: Cag pathogenicity island protein
AX: Type IV secretion system apparatus protein CagX
AY: Cag pathogenicity island protein (Cag7)
BT: Cag pathogenicity island protein
BX: Type IV secretion system apparatus protein CagX
BY: Cag pathogenicity island protein (Cag7)
CT: Cag pathogenicity island protein
CX: Type IV secretion system apparatus protein CagX
CY: Cag pathogenicity island protein (Cag7)
DT: Cag pathogenicity island protein
DX: Type IV secretion system apparatus protein CagX
DY: Cag pathogenicity island protein (Cag7)
ET: Cag pathogenicity island protein
EX: Type IV secretion system apparatus protein CagX
EY: Cag pathogenicity island protein (Cag7)
FT: Cag pathogenicity island protein
FX: Type IV secretion system apparatus protein CagX
FY: Cag pathogenicity island protein (Cag7)
GT: Cag pathogenicity island protein
GX: Type IV secretion system apparatus protein CagX
GY: Cag pathogenicity island protein (Cag7)
HT: Cag pathogenicity island protein
HX: Type IV secretion system apparatus protein CagX
HY: Cag pathogenicity island protein (Cag7)
IT: Cag pathogenicity island protein
IX: Type IV secretion system apparatus protein CagX
IY: Cag pathogenicity island protein (Cag7)
JT: Cag pathogenicity island protein
JX: Type IV secretion system apparatus protein CagX
JY: Cag pathogenicity island protein (Cag7)
KT: Cag pathogenicity island protein
KX: Type IV secretion system apparatus protein CagX
KY: Cag pathogenicity island protein (Cag7)
LT: Cag pathogenicity island protein
LX: Type IV secretion system apparatus protein CagX
LY: Cag pathogenicity island protein (Cag7)
MT: Cag pathogenicity island protein
MX: Type IV secretion system apparatus protein CagX
MY: Cag pathogenicity island protein (Cag7)
NT: Cag pathogenicity island protein
NX: Type IV secretion system apparatus protein CagX
NY: Cag pathogenicity island protein (Cag7)


Theoretical massNumber of molelcules
Total (without water)4,377,82642
Polymers4,377,82642
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area249180 Å2
ΔGint-1176 kcal/mol
Surface area309570 Å2
Noncrystallographic symmetry (NCS)NCS domain:
IDEns-IDDetails
d_1ens_1chain "DT"
d_2ens_1chain "BT"
d_3ens_1chain "CT"
d_4ens_1chain "IT"
d_5ens_1chain "ET"
d_6ens_1chain "FT"
d_7ens_1chain "GT"
d_8ens_1chain "HT"
d_9ens_1chain "KT"
d_10ens_1chain "JT"
d_11ens_1chain "AT"
d_12ens_1chain "LT"
d_13ens_1chain "MT"
d_14ens_1chain "NT"
d_1ens_2chain "DX"
d_2ens_2chain "BX"
d_3ens_2chain "CX"
d_4ens_2chain "IX"
d_5ens_2chain "EX"
d_6ens_2chain "FX"
d_7ens_2chain "GX"
d_8ens_2chain "HX"
d_9ens_2chain "JX"
d_10ens_2chain "AX"
d_11ens_2chain "KX"
d_12ens_2chain "LX"
d_13ens_2chain "MX"
d_14ens_2chain "NX"
d_1ens_3chain "AY"
d_2ens_3chain "BY"
d_3ens_3chain "CY"
d_4ens_3chain "DY"
d_5ens_3chain "EY"
d_6ens_3chain "FY"
d_7ens_3chain "GY"
d_8ens_3chain "HY"
d_9ens_3chain "IY"
d_10ens_3chain "JY"
d_11ens_3chain "KY"
d_12ens_3chain "LY"
d_13ens_3chain "MY"
d_14ens_3chain "NY"

NCS domain segments:
Dom-IDComponent-IDEns-IDBeg label comp-IDEnd label comp-IDLabel asym-IDLabel seq-ID
d_11ens_1LYSSERJ1 - 179
d_21ens_1LYSSERD1 - 179
d_31ens_1LYSSERG1 - 179
d_41ens_1LYSSERY1 - 179
d_51ens_1LYSSERM1 - 179
d_61ens_1LYSSERP1 - 179
d_71ens_1LYSSERS1 - 179
d_81ens_1LYSSERV1 - 179
d_91ens_1LYSSEREA1 - 179
d_101ens_1LYSSERBA1 - 179
d_111ens_1LYSSERA1 - 179
d_121ens_1LYSSERHA1 - 179
d_131ens_1LYSSERKA1 - 179
d_141ens_1LYSSERNA1 - 179
d_11ens_2GLNLYSK1 - 155
d_21ens_2GLNLYSE1 - 155
d_31ens_2GLNLYSH1 - 155
d_41ens_2GLNLYSZ1 - 155
d_51ens_2GLNLYSN1 - 155
d_61ens_2GLNLYSQ1 - 155
d_71ens_2GLNLYST1 - 155
d_81ens_2GLNLYSW1 - 155
d_91ens_2GLNLYSCA1 - 155
d_101ens_2GLNLYSB1 - 155
d_111ens_2GLNLYSFA1 - 155
d_121ens_2GLNLYSIA1 - 155
d_131ens_2GLNLYSLA1 - 155
d_141ens_2GLNLYSOA1 - 155
d_11ens_3PROLYSC1 - 209
d_21ens_3PROLYSF1 - 209
d_31ens_3PROLYSI1 - 209
d_41ens_3PROLYSL1 - 209
d_51ens_3PROLYSO1 - 209
d_61ens_3PROLYSR1 - 209
d_71ens_3PROLYSU1 - 209
d_81ens_3PROLYSX1 - 209
d_91ens_3PROLYSAA1 - 209
d_101ens_3PROLYSDA1 - 209
d_111ens_3PROLYSGA1 - 209
d_121ens_3PROLYSJA1 - 209
d_131ens_3PROLYSMA1 - 209
d_141ens_3PROLYSPA1 - 209

NCS ensembles :
ID
ens_1
ens_2
ens_3

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Components

#1: Protein
Cag pathogenicity island protein / Cag12 / Type IV secretion system apparatus protein CagT


Mass: 32232.738 Da / Num. of mol.: 14 / Source method: isolated from a natural source / Source: (natural) Helicobacter pylori (bacteria) / References: UniProt: Q6VRP0
#2: Protein
Type IV secretion system apparatus protein CagX / Secretion


Mass: 60720.480 Da / Num. of mol.: 14 / Source method: isolated from a natural source / Source: (natural) Helicobacter pylori (bacteria) / References: UniProt: A0A2J9KJM4
#3: Protein
Cag pathogenicity island protein (Cag7)


Mass: 219748.641 Da / Num. of mol.: 14 / Source method: isolated from a natural source / Source: (natural) Helicobacter pylori (bacteria) / References: UniProt: O25262

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Helicobacter pylori dCag3 Cag T4SS OMC / Type: COMPLEX / Entity ID: all / Source: NATURAL
Source (natural)Organism: Helicobacter pylori (bacteria)
Buffer solutionpH: 7
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy
Image recordingElectron dose: 59.7 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.18.2_3874refinement
PHENIX1.18.2_3874refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C14 (14 fold cyclic)
3D reconstructionResolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 7337 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 75.84 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.010961992
ELECTRON MICROSCOPYf_angle_d1.211483594
ELECTRON MICROSCOPYf_chiral_restr0.06959520
ELECTRON MICROSCOPYf_plane_restr0.007310598
ELECTRON MICROSCOPYf_dihedral_angle_d18.17338134
Refine LS restraints NCS
Ens-IDDom-IDAuth asym-IDRefine-IDTypeRms dev position (Å)
ens_1d_2BTELECTRON MICROSCOPYNCS constraints0.0940309561093
ens_1d_3CTELECTRON MICROSCOPYNCS constraints0.000699363941904
ens_1d_4ITELECTRON MICROSCOPYNCS constraints0.0940288872681
ens_1d_5ETELECTRON MICROSCOPYNCS constraints0.0940354865394
ens_1d_6FTELECTRON MICROSCOPYNCS constraints0.0940406120581
ens_1d_7GTELECTRON MICROSCOPYNCS constraints0.000704816890503
ens_1d_8HTELECTRON MICROSCOPYNCS constraints0.0940381475833
ens_1d_9KTELECTRON MICROSCOPYNCS constraints0.000710887873727
ens_1d_10JTELECTRON MICROSCOPYNCS constraints0.000696709803386
ens_1d_11ATELECTRON MICROSCOPYNCS constraints0.0940362342342
ens_1d_12LTELECTRON MICROSCOPYNCS constraints0.000698362008439
ens_1d_13MTELECTRON MICROSCOPYNCS constraints0.0940506006959
ens_1d_14NTELECTRON MICROSCOPYNCS constraints0.0940374615458
ens_2d_2BXELECTRON MICROSCOPYNCS constraints0.000701254680986
ens_2d_3CXELECTRON MICROSCOPYNCS constraints0.102150979939
ens_2d_4IXELECTRON MICROSCOPYNCS constraints0.102142489231
ens_2d_5EXELECTRON MICROSCOPYNCS constraints0.000704160413336
ens_2d_6FXELECTRON MICROSCOPYNCS constraints0.000705101633799
ens_2d_7GXELECTRON MICROSCOPYNCS constraints0.000711113206891
ens_2d_8HXELECTRON MICROSCOPYNCS constraints0.000700970298253
ens_2d_9JXELECTRON MICROSCOPYNCS constraints0.10214395112
ens_2d_10AXELECTRON MICROSCOPYNCS constraints0.1021407285
ens_2d_11KXELECTRON MICROSCOPYNCS constraints0.000707803301165
ens_2d_12LXELECTRON MICROSCOPYNCS constraints0.000697950776351
ens_2d_13MXELECTRON MICROSCOPYNCS constraints0.102137238921
ens_2d_14NXELECTRON MICROSCOPYNCS constraints0.000704150099729
ens_3d_2BYELECTRON MICROSCOPYNCS constraints0.000709143587492
ens_3d_3CYELECTRON MICROSCOPYNCS constraints0.000702502147747
ens_3d_4DYELECTRON MICROSCOPYNCS constraints0.000705171815389
ens_3d_5EYELECTRON MICROSCOPYNCS constraints0.000712090006236
ens_3d_6FYELECTRON MICROSCOPYNCS constraints0.000703203130789
ens_3d_7GYELECTRON MICROSCOPYNCS constraints0.000721915829642
ens_3d_8HYELECTRON MICROSCOPYNCS constraints0.00071168623045
ens_3d_9IYELECTRON MICROSCOPYNCS constraints0.000715995548635
ens_3d_10JYELECTRON MICROSCOPYNCS constraints0.000708237685559
ens_3d_11KYELECTRON MICROSCOPYNCS constraints0.00071371756982
ens_3d_12LYELECTRON MICROSCOPYNCS constraints0.000703368379152
ens_3d_13MYELECTRON MICROSCOPYNCS constraints0.000711466874551
ens_3d_14NYELECTRON MICROSCOPYNCS constraints0.000712745306183

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