+Open data
-Basic information
Entry | Database: PDB / ID: 7nri | ||||||
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Title | Structure of the darobactin-bound E. coli BAM complex (BamABCDE) | ||||||
Components |
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Keywords | MEMBRANE PROTEIN / Beta-Barrel / outer membrane / protein insertion / protein folding / protein maturation / antibiotic / natural product / cyclized peptide | ||||||
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 / membrane / identical protein binding Similarity search - Function | ||||||
Biological species | Escherichia coli (E. coli) synthetic construct (others) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.03 Å | ||||||
Authors | Jakob, R.P. / Kaur, H. / Marzinek, J.K. / Green, R. / Imai, Y. / Bolla, J. / Robinson, C. / Bond, P.J. / Lewis, K. / Maier, T. / Hiller, S. | ||||||
Citation | Journal: Nature / Year: 2021 Title: The antibiotic darobactin mimics a β-strand to inhibit outer membrane insertase. Authors: Hundeep Kaur / Roman P Jakob / Jan K Marzinek / Robert Green / Yu Imai / Jani Reddy Bolla / Elia Agustoni / Carol V Robinson / Peter J Bond / Kim Lewis / Timm Maier / Sebastian Hiller / Abstract: Antibiotics that target Gram-negative bacteria in new ways are needed to resolve the antimicrobial resistance crisis. Gram-negative bacteria are protected by an additional outer membrane, rendering ...Antibiotics that target Gram-negative bacteria in new ways are needed to resolve the antimicrobial resistance crisis. Gram-negative bacteria are protected by an additional outer membrane, rendering proteins on the cell surface attractive drug targets. The natural compound darobactin targets the bacterial insertase BamA-the central unit of the essential BAM complex, which facilitates the folding and insertion of outer membrane proteins. BamA lacks a typical catalytic centre, and it is not obvious how a small molecule such as darobactin might inhibit its function. Here we resolve the mode of action of darobactin at the atomic level using a combination of cryo-electron microscopy, X-ray crystallography, native mass spectrometry, in vivo experiments and molecular dynamics simulations. Two cyclizations pre-organize the darobactin peptide in a rigid β-strand conformation. This creates a mimic of the recognition signal of native substrates with a superior ability to bind to the lateral gate of BamA. Upon binding, darobactin replaces a lipid molecule from the lateral gate to use the membrane environment as an extended binding pocket. Because the interaction between darobactin and BamA is largely mediated by backbone contacts, it is particularly robust against potential resistance mutations. Our results identify the lateral gate as a functional hotspot in BamA and will allow the rational design of antibiotics that target this bacterial Achilles heel. | ||||||
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 | 7nri.cif.gz | 488.3 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7nri.ent.gz | 411.1 KB | Display | PDB format |
PDBx/mmJSON format | 7nri.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/nr/7nri ftp://data.pdbj.org/pub/pdb/validation_reports/nr/7nri | HTTPS FTP |
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-Related structure data
Related structure data | 12546MC 7nreC 7nrfC 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
-Outer membrane protein assembly factor ... , 5 types, 5 molecules ABCDE
#1: Protein | Mass: 88227.430 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (strain K12) (bacteria) Strain: K12 / Gene: bamA, yaeT, yzzN, yzzY, b0177, JW0172 / Plasmid: pQLink / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21(DE3) / Variant (production host): C41 / References: UniProt: P0A940 |
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#2: Protein | Mass: 40924.516 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (strain K12) (bacteria) Strain: K12 / Gene: bamB, yfgL, b2512, JW2496 / Plasmid: pQlink / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21(DE3) / Variant (production host): C41 / References: UniProt: P77774 |
#3: Protein | Mass: 34401.250 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (strain K12) (bacteria) Strain: K12 / Gene: bamC, dapX, nlpB, b2477, JW2462 / Plasmid: pQlink / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21(DE3) / Variant (production host): C41 / References: UniProt: P0A903 |
#4: Protein | Mass: 25816.818 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (strain K12) (bacteria) Strain: K12 / Gene: bamD, yfiO, b2595, JW2577 / Plasmid: pQLink / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21(DE3) / Variant (production host): C41 / References: UniProt: P0AC02 |
#5: Protein | Mass: 11462.772 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (strain K12) (bacteria) Strain: K12 / Gene: bamE, smpA, b2617, JW2598 / Plasmid: pQLink / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21(DE3) / Variant (production host): C41 / References: UniProt: P0A937 |
-Protein/peptide , 1 types, 1 molecules G
#6: Protein/peptide | |
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-Details
Has ligand of interest | Y |
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Nonpolymer details | Original Refinement was done with Darobactin defined as one ligand |
-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: BamABCDE complex / Type: COMPLEX / Entity ID: #2-#5 / Source: RECOMBINANT | ||||||||||||||||||||
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Molecular weight | Experimental value: NO | ||||||||||||||||||||
Source (natural) | Organism: Escherichia coli K-12 (bacteria) | ||||||||||||||||||||
Source (recombinant) | Organism: Escherichia coli BL21(DE3) (bacteria) / Strain: C41 / Plasmid: pQLink | ||||||||||||||||||||
Buffer solution | pH: 8 | ||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 5.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||
Specimen support | Grid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: C-flat-1.2/1.3 | ||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 293 K / Details: 4microL of sample applied |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS / Details: Manually grid screening, operated by SerialEM |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 165000 X / Nominal defocus max: 1800 nm / Nominal defocus min: 800 nm / Calibrated defocus min: 500 nm / Calibrated defocus max: 2200 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 9 sec. / Electron dose: 50 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 5527 |
EM imaging optics | Energyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV |
Image scans | Movie frames/image: 45 |
-Processing
Software | Name: PHENIX / Version: 1.17.1_3660: / Classification: refinement | ||||||||||||||||||||||||||||||
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EM software |
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CTF correction | Type: NONE | ||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 870023 | ||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.03 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 157474 / Num. of class averages: 1 / Symmetry type: POINT | ||||||||||||||||||||||||||||||
Atomic model building | Protocol: AB INITIO MODEL / Space: REAL | ||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 5D0O | ||||||||||||||||||||||||||||||
Refine LS restraints |
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