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- PDB-8p53: Cryo-EM structure of the c-di-GMP-free FleQ-FleN master regulator... -
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Open data
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Basic information
Entry | Database: PDB / ID: 8p53 | ||||||
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Title | Cryo-EM structure of the c-di-GMP-free FleQ-FleN master regulator complex of P. aeruginosa | ||||||
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![]() | GENE REGULATION / Pseudomonas aeruginosa / biofilm / c-di-GMP / transcription regulation | ||||||
Function / homology | ![]() positive regulation of cilium-dependent cell motility / regulation of bacterial-type flagellum-dependent cell motility / negative regulation of cell division / cyclic-di-GMP binding / positive regulation of cell-substrate adhesion / negative regulation of extracellular matrix assembly / DNA-binding transcription repressor activity / DNA-binding transcription activator activity / cis-regulatory region sequence-specific DNA binding / protein-DNA complex ...positive regulation of cilium-dependent cell motility / regulation of bacterial-type flagellum-dependent cell motility / negative regulation of cell division / cyclic-di-GMP binding / positive regulation of cell-substrate adhesion / negative regulation of extracellular matrix assembly / DNA-binding transcription repressor activity / DNA-binding transcription activator activity / cis-regulatory region sequence-specific DNA binding / protein-DNA complex / cytoplasmic side of plasma membrane / transcription cis-regulatory region binding / DNA-templated transcription / regulation of DNA-templated transcription / ATP hydrolysis activity / ATP binding / cytosol Similarity search - Function | ||||||
Biological species | ![]() | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.7 Å | ||||||
![]() | Torres-Sanchez, L. / Krasteva, P.V. | ||||||
Funding support | European Union, 1items
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![]() | ![]() Title: Structures of the FleQ-FleN master regulators reveal large-scale conformational switching in motility and biofilm control. Authors: Lucía Torres-Sánchez / Thibault Géry Sana / Marion Decossas / Yaser Hashem / Petya Violinova Krasteva / ![]() Abstract: can cause a wide array of chronic and acute infections associated with its ability to rapidly switch between planktonic, biofilm, and dispersed lifestyles, each with a specific arsenal for bacterial ... can cause a wide array of chronic and acute infections associated with its ability to rapidly switch between planktonic, biofilm, and dispersed lifestyles, each with a specific arsenal for bacterial survival and virulence. At the cellular level, many of the physiological transitions are orchestrated by the intracellular second messenger c-di-GMP and its receptor-effector FleQ. A bacterial enhancer binding protein, FleQ acts as a master regulator of both flagellar motility and adherence factor secretion and uses remarkably different transcription activation mechanisms depending on its dinucleotide loading state, adenosine triphosphatase (ATPase) activity, interactions with polymerase sigma (σ) factors, and complexation with a second ATPase, FleN. How the FleQ-FleN tandem can exert diverse effects through recognition of a conserved FleQ binding consensus has remained enigmatic. Here, we provide cryogenic electron microscopy (cryo-EM) structures of both c-di-GMP-bound and c-di-GMP-free FleQ-FleN complexes which deepen our understanding of the proteins' (di)nucleotide-dependent conformational switching and fine-tuned roles in gene expression regulation. | ||||||
History |
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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PDBx/mmCIF format | ![]() | 408.6 KB | Display | ![]() |
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PDB format | ![]() | 328.1 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
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-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 17445MC ![]() 8pb9C M: map data used to model this data C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
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Assembly
Deposited unit | ![]()
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Components
#1: Protein | Mass: 33236.332 Da / Num. of mol.: 2 / Mutation: FleN D48A Source method: isolated from a genetically manipulated source Details: P. aeruginosa FleN D48A / Source: (gene. exp.) ![]() ![]() ![]() #2: Protein | Mass: 55494.125 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Details: ...Details: MGSWRETKLLLIDDNLDRSRDLAVILNFLGEDQLTCNSEDWREVAAGLSNSREALCVLLGSVESKGGAVELLKQLASWDEYLPILLIGEPAPADWPEELRRRVLASLEMPPSYNKLLDSLHRAQVYREMYDQARERGRSREPNLFRSLVGTSRAIQQVRQMMQQVADTDASVLILGESGTGKEVVARNLHYHSKRREGPFVPVNCGAIPAELLESELFGHEKGAFTGAITSRAGRFELANGGTLFLDEIGDMPLPMQVKLLRVLQERTFERVGSNKTQNVDVRIIAATHKNLEKMIEDGTFREDLYYRLNVFPIEMAPLRERVEDIALLLNELISRMEHEKRGSIRFNSAAIMSLCRHDWPGNVRELANLVERLAIMHPYGVIGVGELPKKFRHVDDEDEQLASSLREELEERAAINAGLPGMDAPAMLPAEGLDLKDYLANLEQGLIQQALDDAGGVVARAAERLRIRRTTLVEKMRKYGMSRRDDDLSDD Source: (gene. exp.) ![]() ![]() ![]() #3: Chemical | #4: Chemical | Has ligand of interest | Y | |
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-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: C-di-GMP-free FleQ-FleN complex / Type: COMPLEX Details: Full-length FleQ co-purified with His-tagged FleN D48A Entity ID: #1-#2 / Source: RECOMBINANT |
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Molecular weight | Value: 0.288 MDa / Experimental value: NO |
Source (natural) | Organism: ![]() |
Source (recombinant) | Organism: ![]() ![]() |
Buffer solution | pH: 8 Details: 20 mM HEPES pH 8.0, 250mM NaCl, 2mM MgCl2, and 2% glycerol |
Specimen | Conc.: 2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Grid material: GOLD / Grid type: UltrAuFoil R1.2/1.3 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 4 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 defocus max: 2500 nm / Nominal defocus min: 300 nm / Cs: 2.7 mm |
Specimen holder | Cryogen: NITROGEN |
Image recording | Electron dose: 51.5 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 21233 / Details: 17099 micrographs retained for processing |
EM imaging optics | Energyfilter name: GIF Quantum LS |
Image scans | Width: 5760 / Height: 4092 |
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Processing
EM software |
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CTF correction | Details: Gctf software / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
Particle selection | Num. of particles selected: 4124200 / Details: 404 228 particles retained for refinement | ||||||||||||||||||||||||
Symmetry | Point symmetry: C2 (2 fold cyclic) | ||||||||||||||||||||||||
3D reconstruction | Resolution: 2.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 404228 / Symmetry type: POINT | ||||||||||||||||||||||||
Atomic model building | Protocol: AB INITIO MODEL / Space: REAL | ||||||||||||||||||||||||
Refine LS restraints |
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