+Open data
-Basic information
Entry | Database: PDB / ID: 5ljo | |||||||||||||||||||||||||||
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Title | E. coli BAM complex (BamABCDE) by cryoEM | |||||||||||||||||||||||||||
Components |
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Keywords | MEMBRANE PROTEIN / BAM / OMP / Beta barrel / Outer membrane / Gram negative | |||||||||||||||||||||||||||
Function / homology | Function and homology information Bam protein complex / Gram-negative-bacterium-type cell outer membrane assembly / protein insertion into membrane / cell outer membrane / protein-macromolecule adaptor activity / cell adhesion / response to antibiotic / cell surface / identical protein binding / membrane Similarity search - Function | |||||||||||||||||||||||||||
Biological species | Escherichia coli K12 (bacteria) Escherichia coli (E. coli) | |||||||||||||||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.9 Å | |||||||||||||||||||||||||||
Authors | Iadanza, M.G. / Ranson, N.A. / Radford, S.E. / Higgins, A.J. / Schffrin, B. / Calabrese, A.N. / Ashcroft, A.E. / Brockwell, D.J. | |||||||||||||||||||||||||||
Funding support | United Kingdom, 8items
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Citation | Journal: Nat Commun / Year: 2016 Title: Lateral opening in the intact β-barrel assembly machinery captured by cryo-EM. Authors: Matthew G Iadanza / Anna J Higgins / Bob Schiffrin / Antonio N Calabrese / David J Brockwell / Alison E Ashcroft / Sheena E Radford / Neil A Ranson / Abstract: The β-barrel assembly machinery (BAM) is a ∼203 kDa complex of five proteins (BamA-E), which is essential for viability in E. coli. BAM promotes the folding and insertion of β-barrel proteins ...The β-barrel assembly machinery (BAM) is a ∼203 kDa complex of five proteins (BamA-E), which is essential for viability in E. coli. BAM promotes the folding and insertion of β-barrel proteins into the outer membrane via a poorly understood mechanism. Several current models suggest that BAM functions through a 'lateral gating' motion of the β-barrel of BamA. Here we present a cryo-EM structure of the BamABCDE complex, at 4.9 Å resolution. The structure is in a laterally open conformation showing that gating is independent of BamB binding. We describe conformational changes throughout the complex and interactions between BamA, B, D and E, and the detergent micelle that suggest communication between BAM and the lipid bilayer. Finally, using an enhanced reconstitution protocol and functional assays, we show that for the outer membrane protein OmpT, efficient folding in vitro requires lateral gating in BAM. | |||||||||||||||||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 5ljo.cif.gz | 537.4 KB | Display | PDBx/mmCIF format |
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PDB format | pdb5ljo.ent.gz | 425.9 KB | Display | PDB format |
PDBx/mmJSON format | 5ljo.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 5ljo_validation.pdf.gz | 775.4 KB | Display | wwPDB validaton report |
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Full document | 5ljo_full_validation.pdf.gz | 790.9 KB | Display | |
Data in XML | 5ljo_validation.xml.gz | 52.1 KB | Display | |
Data in CIF | 5ljo_validation.cif.gz | 79.7 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/lj/5ljo ftp://data.pdbj.org/pub/pdb/validation_reports/lj/5ljo | HTTPS FTP |
-Related structure data
Related structure data | 4061MC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 39692.156 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli K12 (bacteria) / Gene: bamB, yfgL, b2512, JW2496 / Production host: Escherichia coli (E. coli) / References: UniProt: P77774 |
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#2: Protein | Mass: 17858.889 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli K12 (bacteria) / Gene: bamC, dapX, nlpB, b2477, JW2462 / Production host: Escherichia coli (E. coli) / References: UniProt: P0A903 |
#3: Protein | Mass: 25008.967 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) / Gene: bamD, yfiO, Z3889, ECs3458 / Production host: Escherichia coli (E. coli) / References: UniProt: P0AC04, UniProt: P0AC02*PLUS |
#4: Protein | Mass: 9728.837 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) / Gene: bamE, smpA, c3139 / Production host: Escherichia coli (E. coli) / References: UniProt: P0A938, UniProt: P0A937*PLUS |
#5: Protein | Mass: 87783.945 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) / Gene: bamA, yaeT, ECS88_0187 / Production host: Escherichia coli (E. coli) / References: UniProt: B7MBF8, UniProt: P0A940*PLUS |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: BAM complex / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT | ||||||||||||||||||||
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Molecular weight | Value: 0.2 MDa / Experimental value: YES | ||||||||||||||||||||
Source (natural) | Organism: Escherichia coli (E. coli) | ||||||||||||||||||||
Source (recombinant) | Organism: Escherichia coli (E. coli) / Plasmid: pJH114 | ||||||||||||||||||||
Buffer solution | pH: 8 | ||||||||||||||||||||
Buffer component |
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Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||
Specimen support | Details: Pelco Easyglo / Grid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: Quantifoil R3.5/1 | ||||||||||||||||||||
Vitrification | Instrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 277 K Details: Double blot. 3 ul sampel applied and blotted by hand, then additional 3 ul sample applied, blotted, and plunge frozen |
-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: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 4000 nm / Nominal defocus min: 2000 nm / Calibrated defocus min: 1500 nm / Calibrated defocus max: 3500 nm / Cs: 2.6 mm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 10 sec. / Electron dose: 40 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 |
Image scans | Sampling size: 5 µm / Width: 3480 / Height: 3712 / Movie frames/image: 20 / Used frames/image: 4-14 |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 472857 | |||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 4.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 95878 / Algorithm: BACK PROJECTION / Symmetry type: POINT | |||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: FLEXIBLE FIT / Space: REAL |