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- PDB-8bo2: BAM-EspP complex structure with BamA-S425C/EspP-S1299C mutations ... -

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Basic information

Entry
Database: PDB / ID: 8bo2
TitleBAM-EspP complex structure with BamA-S425C/EspP-S1299C mutations in nanodisc
Components
  • (Outer membrane protein assembly factor ...) x 5
  • Serine protease EspP
KeywordsTRANSPORT PROTEIN / BAM / BamABCDE / EspP / Gram-negative bacteria / outer membrane protein / outer membrane barrel / BamA / BamB / BamC / BamD / BamE / MEMBRANE PROTEIN / Outer membrane protein insertion and release
Function / homology
Function and homology information


Bam protein complex / Gram-negative-bacterium-type cell outer membrane assembly / protein insertion into membrane / Hydrolases; Acting on peptide bonds (peptidases); Serine endopeptidases / cell outer membrane / protein-macromolecule adaptor activity / periplasmic space / serine-type endopeptidase activity / response to antibiotic / cell surface ...Bam protein complex / Gram-negative-bacterium-type cell outer membrane assembly / protein insertion into membrane / Hydrolases; Acting on peptide bonds (peptidases); Serine endopeptidases / cell outer membrane / protein-macromolecule adaptor activity / periplasmic space / serine-type endopeptidase activity / response to antibiotic / cell surface / proteolysis / extracellular region / identical protein binding / membrane
Similarity search - Function
membrane protein fhac: a member of the omp85/tpsb transporter family / membrane protein fhac / Peptidase S6, IgA endopeptidase / Peptidase family S6 domain / Immunoglobulin A1 protease / Peptidase family S6 domain profile. / Autotransporter beta-domain / Outer membrane autotransporter barrel / Autotransporter beta-domain / Autotransporter beta-domain profile. ...membrane protein fhac: a member of the omp85/tpsb transporter family / membrane protein fhac / Peptidase S6, IgA endopeptidase / Peptidase family S6 domain / Immunoglobulin A1 protease / Peptidase family S6 domain profile. / Autotransporter beta-domain / Outer membrane autotransporter barrel / Autotransporter beta-domain / Autotransporter beta-domain profile. / Autotransporter beta-domain / Autotransporter beta-domain superfamily / Autotransporter, pectate lyase C-like domain superfamily / Outer membrane protein assembly factor BamB / Outer membrane protein assembly factor BamC / NlpB/DapX lipoprotein / Outer membrane protein assembly factor BamC, C-terminal / NlpB/DapX lipoprotein / Outer membrane protein assembly factor BamB / Outer membrane protein assembly factor BamE / Lipoprotein SmpA/OmlA / Outer membrane protein assembly factor BamE domain / Outer membrane protein assembly factor BamD / Outer membrane lipoprotein BamD-like / Outer membrane lipoprotein / BamE-like / Pyrrolo-quinoline quinone repeat / Outer membrane protein assembly factor BamA / POTRA domain, BamA/TamA-like / Surface antigen variable number repeat / Surface antigen D15-like / Pyrrolo-quinoline quinone beta-propeller repeat / beta-propeller repeat / POTRA domain / POTRA domain profile. / Bacterial surface antigen (D15) / Omp85 superfamily domain / Porin / Pectin lyase fold/virulence factor / Quinoprotein alcohol dehydrogenase-like superfamily / Ubiquitin-like (UB roll) / Prokaryotic membrane lipoprotein lipid attachment site profile. / Tetratricopeptide-like helical domain superfamily / WD40/YVTN repeat-like-containing domain superfamily / Roll / Beta Barrel / Mainly Beta / Alpha Beta
Similarity search - Domain/homology
Outer membrane protein assembly factor BamA / Outer membrane protein assembly factor BamC / Outer membrane protein assembly factor BamE / Outer membrane protein assembly factor BamD / Outer membrane protein assembly factor BamB / Serine protease EspP
Similarity search - Component
Biological speciesEscherichia coli K-12 (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsShen, C. / Chang, S. / Luo, Q. / Zhang, Z. / Xie, T. / Luo, B. / Lu, G. / Zhu, X. / Wei, X. / Dong, C. ...Shen, C. / Chang, S. / Luo, Q. / Zhang, Z. / Xie, T. / Luo, B. / Lu, G. / Zhu, X. / Wei, X. / Dong, C. / Zhou, R. / Zhang, X. / Tang, X. / Dong, H.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)31900039,32000844,81971974 China
CitationJournal: Nature / Year: 2023
Title: Structural basis of BAM-mediated outer membrane β-barrel protein assembly.
Authors: Chongrong Shen / Shenghai Chang / Qinghua Luo / Kevin Chun Chan / Zhibo Zhang / Bingnan Luo / Teng Xie / Guangwen Lu / Xiaofeng Zhu / Xiawei Wei / Changjiang Dong / Ruhong Zhou / Xing Zhang ...Authors: Chongrong Shen / Shenghai Chang / Qinghua Luo / Kevin Chun Chan / Zhibo Zhang / Bingnan Luo / Teng Xie / Guangwen Lu / Xiaofeng Zhu / Xiawei Wei / Changjiang Dong / Ruhong Zhou / Xing Zhang / Xiaodi Tang / Haohao Dong /
Abstract: The outer membrane structure is common in Gram-negative bacteria, mitochondria and chloroplasts, and contains outer membrane β-barrel proteins (OMPs) that are essential interchange portals of ...The outer membrane structure is common in Gram-negative bacteria, mitochondria and chloroplasts, and contains outer membrane β-barrel proteins (OMPs) that are essential interchange portals of materials. All known OMPs share the antiparallel β-strand topology, implicating a common evolutionary origin and conserved folding mechanism. Models have been proposed for bacterial β-barrel assembly machinery (BAM) to initiate OMP folding; however, mechanisms by which BAM proceeds to complete OMP assembly remain unclear. Here we report intermediate structures of BAM assembling an OMP substrate, EspP, demonstrating sequential conformational dynamics of BAM during the late stages of OMP assembly, which is further supported by molecular dynamics simulations. Mutagenic in vitro and in vivo assembly assays reveal functional residues of BamA and EspP for barrel hybridization, closure and release. Our work provides novel insights into the common mechanism of OMP assembly.
History
DepositionNov 14, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0Apr 26, 2023Provider: repository / Type: Initial release
Revision 1.1May 10, 2023Group: Database references / Category: citation / citation_author / Item: _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.2May 17, 2023Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.3May 24, 2023Group: Database references / Category: citation
Item: _citation.page_first / _citation.page_last ..._citation.page_first / _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.4Nov 13, 2024Group: Data collection / Structure summary
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Outer membrane protein assembly factor BamA
B: Outer membrane protein assembly factor BamB
C: Outer membrane protein assembly factor BamC
D: Outer membrane protein assembly factor BamD
E: Outer membrane protein assembly factor BamE
P: Serine protease EspP


Theoretical massNumber of molelcules
Total (without water)247,2116
Polymers247,2116
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area15190 Å2
ΔGint-73 kcal/mol
Surface area76150 Å2
MethodPISA

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Components

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Outer membrane protein assembly factor ... , 5 types, 5 molecules ABCDE

#1: Protein Outer membrane protein assembly factor BamA


Mass: 88273.555 Da / Num. of mol.: 1 / Mutation: S425C
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Strain: K12 / DH10B / Gene: bamA, yaeT, ECDH10B_0157
Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
References: UniProt: B1XD46
#2: Protein Outer membrane protein assembly factor BamB


Mass: 41918.945 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: bamB, yfgL, b2512, JW2496
Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
References: UniProt: P77774
#3: Protein Outer membrane protein assembly factor BamC


Mass: 36875.277 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: bamC, dapX, nlpB, b2477, JW2462
Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
References: UniProt: P0A903
#4: Protein Outer membrane protein assembly factor BamD


Mass: 27858.350 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: bamD, yfiO, b2595, JW2577
Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
References: UniProt: P0AC02
#5: Protein Outer membrane protein assembly factor BamE


Mass: 13530.256 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: bamE, smpA, b2617, JW2598
Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
References: UniProt: P0A937

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Protein , 1 types, 1 molecules P

#6: Protein Serine protease EspP


Mass: 38754.836 Da / Num. of mol.: 1 / Mutation: S1299C
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: espP, L7020, ECO57PM78
Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
References: UniProt: Q7BSW5, Hydrolases; Acting on peptide bonds (peptidases); Serine endopeptidases

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Details

Has protein modificationY

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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

Component
IDNameTypeEntity IDParent-IDSource
1BAM-EspP complex with BamA-G431C/EspP-N1293C mutations in nanodiscCOMPLEXall0RECOMBINANT
2BAMCOMPLEX#1-#51RECOMBINANT
3EspPCOMPLEX#61RECOMBINANT
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
21Escherichia coli K-12 (bacteria)83333
32Escherichia coli K-12 (bacteria)83333
43Escherichia coli O157:H7 (bacteria)83334
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
21Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)866768
32Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)866768
43Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)866768
Buffer solutionpH: 7.8
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 FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 1000 nm / Calibrated defocus min: 1000 nm / Calibrated defocus max: 2000 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k)

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.20.1_4487refinement
PHENIX1.20.1_4487refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 203494 / Symmetry type: POINT
RefinementCross valid method: NONE

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