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- PDB-6ye4: Structure of ExbB pentamer from Serratia marcescens by single par... -
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Basic information
Entry | Database: PDB / ID: 6ye4 | ||||||
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Title | Structure of ExbB pentamer from Serratia marcescens by single particle cryo electron microscopy | ||||||
![]() | Biopolymer transport protein ExbB | ||||||
![]() | METAL TRANSPORT / membrane protein / iron uptake / proton transfer / TonB complex | ||||||
Function / homology | TonB-system energizer ExbB type-1 / MotA/TolQ/ExbB proton channel / MotA/TolQ/ExbB proton channel family / transmembrane transporter activity / protein transport / membrane => GO:0016020 / plasma membrane / Chem-PGT / Biopolymer transport protein ExbB![]() | ||||||
Biological species | ![]() | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å | ||||||
![]() | Biou, V. / Delepelaire, P. / Coureux, P.D. / Chami, M. | ||||||
Funding support | ![]()
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![]() | ![]() Title: Structural and molecular determinants for the interaction of ExbB from Serratia marcescens and HasB, a TonB paralog. Authors: Valérie Biou / Ricardo Jorge Diogo Adaixo / Mohamed Chami / Pierre-Damien Coureux / Benoist Laurent / Véronique Yvette Ntsogo Enguéné / Gisele Cardoso de Amorim / Nadia Izadi-Pruneyre / ...Authors: Valérie Biou / Ricardo Jorge Diogo Adaixo / Mohamed Chami / Pierre-Damien Coureux / Benoist Laurent / Véronique Yvette Ntsogo Enguéné / Gisele Cardoso de Amorim / Nadia Izadi-Pruneyre / Christian Malosse / Julia Chamot-Rooke / Henning Stahlberg / Philippe Delepelaire / ![]() ![]() ![]() Abstract: ExbB and ExbD are cytoplasmic membrane proteins that associate with TonB to convey the energy of the proton-motive force to outer membrane receptors in Gram-negative bacteria for iron uptake. The ...ExbB and ExbD are cytoplasmic membrane proteins that associate with TonB to convey the energy of the proton-motive force to outer membrane receptors in Gram-negative bacteria for iron uptake. The opportunistic pathogen Serratia marcescens (Sm) possesses both TonB and a heme-specific TonB paralog, HasB. ExbB has a long periplasmic extension absent in other bacteria such as E. coli (Ec). Long ExbB's are found in several genera of Alphaproteobacteria, most often in correlation with a hasB gene. We investigated specificity determinants of ExbB and HasB. We determined the cryo-EM structures of ExbB and of the ExbB-ExbD complex from S. marcescens. ExbB alone is a stable pentamer, and its complex includes two ExbD monomers. We showed that ExbB extension interacts with HasB and is involved in heme acquisition and we identified key residues in the membrane domain of ExbB and ExbB, essential for function and likely involved in the interaction with TonB/HasB. Our results shed light on the class of inner membrane energy machinery formed by ExbB, ExbD and HasB. #1: ![]() Title: Functional and structural characterization of Serratia marcescens ExbB: determinants of the interaction with HasB/TonB Authors: Biou, V. / Chami, M. / Coureux, P.D. / Laurent, B. / Ntsogo, Y. / Izadi-Pruneyre, N. / Malosse, C. / Chamot-Rooke, J. / Stahlberg, H. / Delepelaire, P. #2: Journal: J Struct Biol / Year: 2012 Title: RELION: implementation of a Bayesian approach to cryo-EM structure determination. Authors: Sjors H W Scheres / ![]() Abstract: RELION, for REgularized LIkelihood OptimizatioN, is an open-source computer program for the refinement of macromolecular structures by single-particle analysis of electron cryo-microscopy (cryo-EM) ...RELION, for REgularized LIkelihood OptimizatioN, is an open-source computer program for the refinement of macromolecular structures by single-particle analysis of electron cryo-microscopy (cryo-EM) data. Whereas alternative approaches often rely on user expertise for the tuning of parameters, RELION uses a Bayesian approach to infer parameters of a statistical model from the data. This paper describes developments that reduce the computational costs of the underlying maximum a posteriori (MAP) algorithm, as well as statistical considerations that yield new insights into the accuracy with which the relative orientations of individual particles may be determined. A so-called gold-standard Fourier shell correlation (FSC) procedure to prevent overfitting is also described. The resulting implementation yields high-quality reconstructions and reliable resolution estimates with minimal user intervention and at acceptable computational costs. | ||||||
History |
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Structure visualization
Movie |
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Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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PDBx/mmCIF format | ![]() | 214 KB | Display | ![]() |
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PDB format | ![]() | 168.3 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
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-Validation report
Summary document | ![]() | 1.3 MB | Display | ![]() |
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Full document | ![]() | 1.3 MB | Display | |
Data in XML | ![]() | 52 KB | Display | |
Data in CIF | ![]() | 72.3 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 10789MC ![]() 7ajqC M: map data used to model this data C: citing same article ( |
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Similar structure data |
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Assembly
Deposited unit | ![]()
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Components
#1: Protein | Mass: 30413.621 Da / Num. of mol.: 5 Source method: isolated from a genetically manipulated source Details: a 6-histidine tag is present at the C-terminus / Source: (gene. exp.) ![]() ![]() ![]() #2: Chemical | ChemComp-PGT / ( 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: Homopentamer of ExbB / Type: COMPLEX Details: the expressed sequence corresponds to the mature sequence after signal peptide cleavage. Entity ID: #1 / Source: RECOMBINANT | ||||||||||||||||||||
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Molecular weight | Value: 0.174 MDa / Experimental value: NO | ||||||||||||||||||||
Source (natural) | Organism: ![]() | ||||||||||||||||||||
Source (recombinant) | Organism: ![]() ![]() | ||||||||||||||||||||
Buffer solution | pH: 8 / Details: 20mM Tris-HCl pH 8,0 100mM NaCl 0,0015% LMNG | ||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 0.1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES Details: the sample was monodisperse as evidenced by gel filtration column | ||||||||||||||||||||
Specimen support | Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/4 | ||||||||||||||||||||
Vitrification | Instrument: LEICA EM CPC / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 277 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 magnification: 165000 X / Nominal defocus max: 2500 nm / Nominal defocus min: 1500 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 93 K / Temperature (min): 83 K |
Image recording | Average exposure time: 7 sec. / Electron dose: 55.95 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 3122 Details: frames were weighted according to electron dose and particle movement during Relion bayesian polishing procedure. |
EM imaging optics | Energyfilter name: GIF Quantum LS / Energyfilter slit width: 20 eV |
Image scans | Width: 3710 / Height: 3838 / Movie frames/image: 56 / Used frames/image: 1-56 |
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Processing
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EM software |
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Image processing | Details: images were processed with Motioncor2 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
CTF correction | Type: NONE | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 1291382 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C5 (5 fold cyclic) | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 157111 / Algorithm: FOURIER SPACE / Num. of class averages: 1 / Symmetry type: POINT | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | B value: 55.3 / Protocol: FLEXIBLE FIT / Space: REAL / Target criteria: Correlation coefficient Details: the ExbB sequence from S. marcescens was modeled by homology from the 5SV0 monomer from E. coli using Phyre software and the pentamer was generated using the 5SV0 symmetry. real space ...Details: the ExbB sequence from S. marcescens was modeled by homology from the 5SV0 monomer from E. coli using Phyre software and the pentamer was generated using the 5SV0 symmetry. real space refinement was carried out with rigid body, simulated annealing and morphing steps. | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 5SV0 Pdb chain-ID: A / Accession code: 5SV0 / Pdb chain residue range: 10-234 / Source name: PDB / Type: experimental model | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 45.97 Å2 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
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