+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-21959 | ||||||||||||
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Title | Colicin E1 fragment in nanodisc-embedded TolC | ||||||||||||
Map data | asymmetric map of colicin E1 bound to TolC in nanodisc; cropped, sharpened, and resampled | ||||||||||||
Sample |
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Keywords | antibiotic efflux / bacteriocin / TRANSPORT PROTEIN / ANTIMICROBIAL PROTEIN | ||||||||||||
Function / homology | Function and homology information negative regulation of ion transmembrane transporter activity / MacAB-TolC complex / pore-forming activity / xenobiotic detoxification by transmembrane export across the cell outer membrane / efflux pump complex / enterobactin transport / enterobactin transmembrane transporter activity / periplasmic side of plasma membrane / bile acid transmembrane transporter activity / xenobiotic detoxification by transmembrane export across the plasma membrane ...negative regulation of ion transmembrane transporter activity / MacAB-TolC complex / pore-forming activity / xenobiotic detoxification by transmembrane export across the cell outer membrane / efflux pump complex / enterobactin transport / enterobactin transmembrane transporter activity / periplasmic side of plasma membrane / bile acid transmembrane transporter activity / xenobiotic detoxification by transmembrane export across the plasma membrane / bile acid and bile salt transport / porin activity / efflux transmembrane transporter activity / monoatomic ion channel activity / cell outer membrane / response to organic cyclic compound / response to toxic substance / monoatomic ion transmembrane transport / outer membrane-bounded periplasmic space / defense response to Gram-negative bacterium / transmembrane transporter binding / killing of cells of another organism / response to xenobiotic stimulus / response to antibiotic / identical protein binding / membrane / plasma membrane Similarity search - Function | ||||||||||||
Biological species | Escherichia coli K-12 (bacteria) / Escherichia coli (E. coli) / Escherichia coli (strain K12) (bacteria) | ||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.09 Å | ||||||||||||
Authors | Kaelber JT / Budiardjo SJ | ||||||||||||
Funding support | United States, 3 items
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Citation | Journal: Elife / Year: 2022 Title: Colicin E1 opens its hinge to plug TolC. Authors: S Jimmy Budiardjo / Jacqueline J Stevens / Anna L Calkins / Ayotunde P Ikujuni / Virangika K Wimalasena / Emre Firlar / David A Case / Julie S Biteen / Jason T Kaelber / Joanna S G Slusky / Abstract: The double membrane architecture of Gram-negative bacteria forms a barrier that is impermeable to most extracellular threats. Bacteriocin proteins evolved to exploit the accessible, surface-exposed ...The double membrane architecture of Gram-negative bacteria forms a barrier that is impermeable to most extracellular threats. Bacteriocin proteins evolved to exploit the accessible, surface-exposed proteins embedded in the outer membrane to deliver cytotoxic cargo. Colicin E1 is a bacteriocin produced by, and lethal to, that hijacks the outer membrane proteins (OMPs) TolC and BtuB to enter the cell. Here, we capture the colicin E1 translocation domain inside its membrane receptor, TolC, by high-resolution cryo-electron microscopy to obtain the first reported structure of a bacteriocin bound to TolC. Colicin E1 binds stably to TolC as an open hinge through the TolC pore-an architectural rearrangement from colicin E1's unbound conformation. This binding is stable in live cells as indicated by single-molecule fluorescence microscopy. Finally, colicin E1 fragments binding to TolC plug the channel, inhibiting its native efflux function as an antibiotic efflux pump, and heightening susceptibility to three antibiotic classes. In addition to demonstrating that these protein fragments are useful starting points for developing novel antibiotic potentiators, this method could be expanded to other colicins to inhibit other OMP functions. | ||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_21959.map.gz | 52.7 MB | EMDB map data format | |
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Header (meta data) | emd-21959-v30.xml emd-21959.xml | 24.4 KB 24.4 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_21959_fsc.xml | 15.1 KB | Display | FSC data file |
Images | emd_21959.png | 64.1 KB | ||
Masks | emd_21959_msk_1.map | 282.6 MB | Mask map | |
Filedesc metadata | emd-21959.cif.gz | 7.1 KB | ||
Others | emd_21959_additional_1.map.gz emd_21959_half_map_1.map.gz emd_21959_half_map_2.map.gz | 145.9 MB 262 MB 262 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-21959 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-21959 | HTTPS FTP |
-Validation report
Summary document | emd_21959_validation.pdf.gz | 1.1 MB | Display | EMDB validaton report |
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Full document | emd_21959_full_validation.pdf.gz | 1.1 MB | Display | |
Data in XML | emd_21959_validation.xml.gz | 22.5 KB | Display | |
Data in CIF | emd_21959_validation.cif.gz | 28.5 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-21959 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-21959 | HTTPS FTP |
-Related structure data
Related structure data | 6wxhMC 6wxiC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_21959.map.gz / Format: CCP4 / Size: 67 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | asymmetric map of colicin E1 bound to TolC in nanodisc; cropped, sharpened, and resampled | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.9 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
File | emd_21959_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Additional map: C3-symmetrized refinement of colicin E1 bound to TolC...
File | emd_21959_additional_1.map | ||||||||||||
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Annotation | C3-symmetrized refinement of colicin E1 bound to TolC in nanodisc; sharpened | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: unmasked, unfiltered cryoSPARC half-map B of colicin E1...
File | emd_21959_half_map_1.map | ||||||||||||
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Annotation | unmasked, unfiltered cryoSPARC half-map B of colicin E1 bound to TolC in nanodisc | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: unmasked, unfiltered cryoSPARC half-map A of colicin E1...
File | emd_21959_half_map_2.map | ||||||||||||
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Annotation | unmasked, unfiltered cryoSPARC half-map A of colicin E1 bound to TolC in nanodisc | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Colicin E1 fragment colE1-T inside nanodisc-embedded TolC
Entire | Name: Colicin E1 fragment colE1-T inside nanodisc-embedded TolC |
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Components |
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-Supramolecule #1: Colicin E1 fragment colE1-T inside nanodisc-embedded TolC
Supramolecule | Name: Colicin E1 fragment colE1-T inside nanodisc-embedded TolC type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Molecular weight | Theoretical: 21.44974 KDa |
-Supramolecule #2: Outer membrane protein TolC
Supramolecule | Name: Outer membrane protein TolC / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1 |
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Source (natural) | Organism: Escherichia coli K-12 (bacteria) |
-Supramolecule #3: Colicin E1 fragment colE1-T
Supramolecule | Name: Colicin E1 fragment colE1-T / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2 |
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Source (natural) | Organism: Escherichia coli (E. coli) |
-Macromolecule #1: Outer membrane protein TolC
Macromolecule | Name: Outer membrane protein TolC / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO |
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Source (natural) | Organism: Escherichia coli (strain K12) (bacteria) / Strain: K12 |
Molecular weight | Theoretical: 53.783355 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: MKKLLPILIG LSLSGFSSLS QAENLMQVYQ QARLSNPELR KSAADRDAAF EKINEARSPL LPQLGLGADY TYSNGYRDAN GINSNATSA SLQLTQSIFD MSKWRALTLQ EKAAGIQDVT YQTDQQTLIL NTATAYFNVL NAIDVLSYTQ AQKEAIYRQL D QTTQRFNV ...String: MKKLLPILIG LSLSGFSSLS QAENLMQVYQ QARLSNPELR KSAADRDAAF EKINEARSPL LPQLGLGADY TYSNGYRDAN GINSNATSA SLQLTQSIFD MSKWRALTLQ EKAAGIQDVT YQTDQQTLIL NTATAYFNVL NAIDVLSYTQ AQKEAIYRQL D QTTQRFNV GLVAITDVQN ARAQYDTVLA NEVTARNNLD NAVEQLRQIT GNYYPELAAL NVENFKTDKP QPVNALLKEA EK RNLSLLQ ARLSQDLARE QIRQAQDGHL PTLDLTASTG ISDTSYSGSK TRGAAGTQYD DSNMGQNKVG LSFSLPIYQG GMV NSQVKQ AQYNFVGASE QLESAHRSVV QTVRSSFNNI NASISSINAY KQAVVSAQSS LDAMEAGYSV GTRTIVDVLD ATTT LYNAK QELANARYNY LINQLNIKSA LGTLNEQDLL ALNNALSKPV STNPENVAPQ TPEQNAIADG YAPDSPAPVV QQTSA RTTT SNGHNPFRN UniProtKB: Outer membrane protein TolC |
-Macromolecule #2: Colicin-E1
Macromolecule | Name: Colicin-E1 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 21.491807 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: METAVAYYKD GVPYDDKGQV IITLLNGTPD GSGSGGGGGK GGSKSESSAA IHATAKWSTA QLKKTQAEQA ARAKAAAEAQ AKAKANRDA LTQRLKDIVN EALRHNASRT PSATELAHAN NAAMQAEDER LRLAKAEEKA RKEAEAAEKA FQEAEQRRKE I EREKAETE ...String: METAVAYYKD GVPYDDKGQV IITLLNGTPD GSGSGGGGGK GGSKSESSAA IHATAKWSTA QLKKTQAEQA ARAKAAAEAQ AKAKANRDA LTQRLKDIVN EALRHNASRT PSATELAHAN NAAMQAEDER LRLAKAEEKA RKEAEAAEKA FQEAEQRRKE I EREKAETE RQLKLAEAEE KRLAALSEEA KALEHHHHHH UniProtKB: Colicin-E1 |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 1 mg/mL | |||||||||
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Buffer | pH: 8 Component:
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Grid | Model: UltrAuFoil / Material: GOLD / Mesh: 200 / Support film - Material: GOLD / Support film - topology: HOLEY ARRAY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR | |||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | FEI TALOS ARCTICA |
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Specialist optics | Energy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 3838 pixel / Digitization - Dimensions - Height: 3710 pixel / Number real images: 10644 / Average exposure time: 6.0 sec. / Average electron dose: 35.96 e/Å2 / Details: 5 frames per second |
Electron beam | Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 20.0 µm / Calibrated defocus max: 2.1 µm / Calibrated defocus min: 0.4 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal magnification: 205000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Talos Arctica / Image courtesy: FEI Company |