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- PDB-7nbx: Lateral-open conformation of the lid-locked BAM complex (BamA E43... -
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Open data
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Basic information
Entry | Database: PDB / ID: 7nbx | ||||||||||||||||||||||||
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Title | Lateral-open conformation of the lid-locked BAM complex (BamA E435C S665C, BamBDCE) by cryoEM | ||||||||||||||||||||||||
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![]() | MEMBRANE PROTEIN / Outer membrane protein assembly / beta-barrel / Gram negative bacteria / protein foldase | ||||||||||||||||||||||||
Function / homology | ![]() Bam protein complex / protein insertion into membrane / Gram-negative-bacterium-type cell outer membrane assembly / cell outer membrane / protein-macromolecule adaptor activity / cell adhesion / response to antibiotic / cell surface / identical protein binding / membrane Similarity search - Function | ||||||||||||||||||||||||
Biological species | ![]() ![]() | ||||||||||||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.8 Å | ||||||||||||||||||||||||
![]() | Haysom, S.F. | ||||||||||||||||||||||||
Funding support | ![]() ![]()
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![]() | ![]() Title: The role of membrane destabilisation and protein dynamics in BAM catalysed OMP folding. Authors: Paul White / Samuel F Haysom / Matthew G Iadanza / Anna J Higgins / Jonathan M Machin / James M Whitehouse / Jim E Horne / Bob Schiffrin / Charlotte Carpenter-Platt / Antonio N Calabrese / ...Authors: Paul White / Samuel F Haysom / Matthew G Iadanza / Anna J Higgins / Jonathan M Machin / James M Whitehouse / Jim E Horne / Bob Schiffrin / Charlotte Carpenter-Platt / Antonio N Calabrese / Kelly M Storek / Steven T Rutherford / David J Brockwell / Neil A Ranson / Sheena E Radford / ![]() ![]() Abstract: The folding of β-barrel outer membrane proteins (OMPs) in Gram-negative bacteria is catalysed by the β-barrel assembly machinery (BAM). How lateral opening in the β-barrel of the major subunit ...The folding of β-barrel outer membrane proteins (OMPs) in Gram-negative bacteria is catalysed by the β-barrel assembly machinery (BAM). How lateral opening in the β-barrel of the major subunit BamA assists in OMP folding, and the contribution of membrane disruption to BAM catalysis remain unresolved. Here, we use an anti-BamA monoclonal antibody fragment (Fab1) and two disulphide-crosslinked BAM variants (lid-locked (LL), and POTRA-5-locked (P5L)) to dissect these roles. Despite being lethal in vivo, we show that all complexes catalyse folding in vitro, albeit less efficiently than wild-type BAM. CryoEM reveals that while Fab1 and BAM-P5L trap an open-barrel state, BAM-LL contains a mixture of closed and contorted, partially-open structures. Finally, all three complexes globally destabilise the lipid bilayer, while BamA does not, revealing that the BAM lipoproteins are required for this function. Together the results provide insights into the role of BAM structure and lipid dynamics in OMP folding. | ||||||||||||||||||||||||
History |
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Structure visualization
Movie |
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Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 477.8 KB | Display | ![]() |
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PDB format | ![]() | 404.3 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Summary document | ![]() | 1.2 MB | Display | ![]() |
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Full document | ![]() | 1.2 MB | Display | |
Data in XML | ![]() | 57 KB | Display | |
Data in CIF | ![]() | 84.8 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 12262MC ![]() 7bm5C ![]() 7bnqC ![]() 7ncsC ![]() 7nd0C C: citing same article ( M: map data used to model this data |
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Similar structure data |
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Links
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Assembly
Deposited unit | ![]()
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Components
#1: Protein | Mass: 90601.352 Da / Num. of mol.: 1 / Mutation: E435C S665C C690S C700S Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Strain: K12 / Gene: bamA, yaeT, yzzN, yzzY, b0177, JW0172 / Production host: ![]() ![]() |
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#2: Protein | Mass: 41918.945 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Strain: K12 / Gene: bamB, yfgL, b2512, JW2496 / Production host: ![]() ![]() |
#3: Protein | Mass: 36875.277 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Strain: K12 / Gene: bamC, dapX, nlpB, b2477, JW2462 / Production host: ![]() ![]() |
#4: Protein | Mass: 27858.350 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Strain: K12 / Gene: bamD, yfiO, b2595, JW2577 / Production host: ![]() ![]() |
#5: Protein | Mass: 13530.256 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Strain: K12 / Gene: bamE, smpA, b2617, JW2598 / Production host: ![]() ![]() |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component | Name: beta-barrel assembly machinery (BAM) complex (BamABCDE) Type: COMPLEX / Entity ID: all / Source: RECOMBINANT | ||||||||||||||||||||
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Molecular weight | Value: 0.203 MDa / Experimental value: NO | ||||||||||||||||||||
Source (natural) | Organism: ![]() ![]() | ||||||||||||||||||||
Source (recombinant) | Organism: ![]() ![]() | ||||||||||||||||||||
Buffer solution | pH: 8 | ||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 3.3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||
Specimen support | Details: Grids were glow discharged in a GlowQube Plus for 30s at 60 mA Grid material: COPPER / Grid type: Quantifoil R1.2/1.3 | ||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.15 K |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 130000 X / Nominal defocus max: 3000 nm / Nominal defocus min: 1500 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 49.7 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) |
EM imaging optics | Energyfilter slit width: 20 eV |
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Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 793444 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 4.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 141612 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | B value: 211 / Protocol: FLEXIBLE FIT / Space: REAL Details: Initial local fitting accomplished in Chimera. VMD was then used to setup cMDFF simulations that were run using NAMD. Both 7BNQ and 5LJO were used as initial models, with the best fitting ...Details: Initial local fitting accomplished in Chimera. VMD was then used to setup cMDFF simulations that were run using NAMD. Both 7BNQ and 5LJO were used as initial models, with the best fitting parts of each then combined and flexibly fit again to generate the final conformation. This was then Real Space Refined in Phenix to generate the final structure | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building |
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