+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-16816 | |||||||||
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Title | Structure of TolQR complex from E.coli | |||||||||
Map data | One of maps used in refinement, Z-flipped for model building. | |||||||||
Sample |
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Keywords | TolQ / TolR / Tol-Pal / complex / inner membrane / MEMBRANE PROTEIN | |||||||||
Function / homology | Function and homology information regulation of membrane invagination / cell envelope / bacteriocin transport / protein import / cell division site / transmembrane transporter activity / protein transport / cell cycle / cell division / membrane / plasma membrane Similarity search - Function | |||||||||
Biological species | Escherichia coli (E. coli) / Escherichia coli K-12 (bacteria) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.2 Å | |||||||||
Authors | Webby MN / Kleanthous C / Press CE | |||||||||
Funding support | United Kingdom, European Union, 2 items
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Citation | Journal: Proc Natl Acad Sci U S A / Year: 2023 Title: Tunable force transduction through the cell envelope. Authors: Daniel P Williams-Jones / Melissa N Webby / Cara E Press / Jan M Gradon / Sophie R Armstrong / Joanna Szczepaniak / Colin Kleanthous / Abstract: The outer membrane (OM) of Gram-negative bacteria is not energised and so processes requiring a driving force must connect to energy-transduction systems in the inner membrane (IM). Tol (Tol-Pal) and ...The outer membrane (OM) of Gram-negative bacteria is not energised and so processes requiring a driving force must connect to energy-transduction systems in the inner membrane (IM). Tol (Tol-Pal) and Ton are related, proton motive force- (PMF-) coupled assemblies that stabilise the OM and import essential nutrients, respectively. Both rely on proton-harvesting IM motor (stator) complexes, which are homologues of the flagellar stator unit Mot, to transduce force to the OM through elongated IM force transducer proteins, TolA and TonB, respectively. How PMF-driven motors in the IM generate mechanical work at the OM via force transducers is unknown. Here, using cryoelectron microscopy, we report the 4.3Å structure of the TolQR motor complex. The structure reaffirms the 5:2 stoichiometry seen in Ton and Mot and, with motor subunits related to each other by 10 to 16° rotation, supports rotary motion as the default for these complexes. We probed the mechanism of force transduction to the OM through in vivo assays of chimeric TolA/TonB proteins where sections of their structurally divergent, periplasm-spanning domains were swapped or replaced by an intrinsically disordered sequence. We find that TolA mutants exhibit a spectrum of force output, which is reflected in their respective abilities to both stabilise the OM and import cytotoxic colicins across the OM. Our studies demonstrate that structural rigidity of force transducer proteins, rather than any particular structural form, drives the efficient conversion of PMF-driven rotary motions of 5:2 motor complexes into physiologically relevant force at the OM. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_16816.map.gz | 96.8 MB | EMDB map data format | |
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Header (meta data) | emd-16816-v30.xml emd-16816.xml | 18.5 KB 18.5 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_16816_fsc.xml | 13.8 KB | Display | FSC data file |
Images | emd_16816.png | 51.3 KB | ||
Filedesc metadata | emd-16816.cif.gz | 5.9 KB | ||
Others | emd_16816_additional_1.map.gz emd_16816_half_map_1.map.gz emd_16816_half_map_2.map.gz | 96.9 MB 95.5 MB 95.5 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-16816 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-16816 | HTTPS FTP |
-Related structure data
Related structure data | 8odtMC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_16816.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Annotation | One of maps used in refinement, Z-flipped for model building. | ||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.832 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Additional map: Class one, from 3D classification.
File | emd_16816_additional_1.map | ||||||||||||
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Annotation | Class one, from 3D classification. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map A
File | emd_16816_half_map_1.map | ||||||||||||
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Annotation | Half map A | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map A
File | emd_16816_half_map_2.map | ||||||||||||
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Annotation | Half map A | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : TolQ-TolR 5:2 complex
Entire | Name: TolQ-TolR 5:2 complex |
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Components |
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-Supramolecule #1: TolQ-TolR 5:2 complex
Supramolecule | Name: TolQ-TolR 5:2 complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: Escherichia coli (E. coli) / Strain: K12 |
Molecular weight | Theoretical: 170 KDa |
-Macromolecule #1: Tol-Pal system protein TolQ
Macromolecule | Name: Tol-Pal system protein TolQ / type: protein_or_peptide / ID: 1 / Number of copies: 5 / Enantiomer: LEVO |
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Source (natural) | Organism: Escherichia coli K-12 (bacteria) |
Molecular weight | Theoretical: 25.623662 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: MTDMNILDLF LKASLLVKLI MLILIGFSIA SWAIIIQRTR ILNAAAREAE AFEDKFWSGI ELSRLYQESQ GKRDNLTGSE QIFYSGFKE FVRLHRANSH APEAVVEGAS RAMRISMNRE LENLETHIPF LGTVGSISPY IGLFGTVWGI MHAFIALGAV K QATLQMVA ...String: MTDMNILDLF LKASLLVKLI MLILIGFSIA SWAIIIQRTR ILNAAAREAE AFEDKFWSGI ELSRLYQESQ GKRDNLTGSE QIFYSGFKE FVRLHRANSH APEAVVEGAS RAMRISMNRE LENLETHIPF LGTVGSISPY IGLFGTVWGI MHAFIALGAV K QATLQMVA PGIAEALIAT AIGLFAAIPA VMAYNRLNQR VNKLELNYDN FMEEFTAILH RQAFTVSESN KG UniProtKB: Tol-Pal system protein TolQ |
-Macromolecule #2: Tol-Pal system protein TolR
Macromolecule | Name: Tol-Pal system protein TolR / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: Escherichia coli K-12 (bacteria) |
Molecular weight | Theoretical: 20.600367 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: MARARGRGRR DLKSEINIVP LLDVLLVLLL IFMATAPIIT QSVEVDLPDA TESQAVSSND NPPVIVEVSG IGQYTVVVEK DRLERLPPE QVVAEVSSRF KANPKTVFLI GGAKDVPYDE IIKALNLLHS AGVKSVGLMT QPILEENLYF QGQFGSWSHP Q FEKGGGSG GGSGGGSWSH PQFEKHHHHH H UniProtKB: Tol-Pal system protein TolR |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 7.5 mg/mL |
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Buffer | pH: 7.5 |
Grid | Model: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 15.0 kPa |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 281 K / Instrument: FEI VITROBOT MARK IV / Details: Blot force 10 Blot time 3 sec Wait time 2 sec. |
Details | sample purified by affinity chromatography (His-tagged TolR) and SEC before application to grids in 50 mM Tris/HCl pH 7.5, 300 mM NaCl, 2 mM EDTA, 0.01% LMNG |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm |
Image recording | Film or detector model: GATAN K3 (6k x 4k) / Average electron dose: 46.39 e/Å2 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |