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- EMDB-10789: Structure of ExbB pentamer from Serratia marcescens by single par... -

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Basic information

Entry
Database: EMDB / ID: EMD-10789
TitleStructure of ExbB pentamer from Serratia marcescens by single particle cryo electron microscopy
Map data
Sample
  • Complex: Homopentamer of ExbB
    • Protein or peptide: Biopolymer transport protein ExbB
  • Ligand: (1S)-2-{[{[(2R)-2,3-DIHYDROXYPROPYL]OXY}(HYDROXY)PHOSPHORYL]OXY}-1-[(PALMITOYLOXY)METHYL]ETHYL STEARATE
Keywordsmembrane protein / iron uptake / proton transfer / TonB complex / METAL TRANSPORT
Function / homologyTonB-system energizer ExbB type-1 / : / MotA/TolQ/ExbB proton channel / MotA/TolQ/ExbB proton channel family / protein import / transmembrane transporter activity / plasma membrane / Biopolymer transport protein ExbB
Function and homology information
Biological speciesSerratia marcescens (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsBiou V / Delepelaire P
Funding support France, 1 items
OrganizationGrant numberCountry
French National Research AgencyANR-11-LABEX-0011-01 France
Citation
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.
#1: Journal: Biorxiv / Year: 2021
Title: Functional and structural characterization of Serratia marcescens ExbB: determinants of the interaction with HasB/TonB
Authors: Biou V / Chami M / Coureux PD / Laurent B / Ntsogo Y / Izadi-Pruneyre N / Malosse C / Chamot-Rooke J / Stahlberg H / Delepelaire P
History
DepositionMar 24, 2020-
Header (metadata) releaseMar 31, 2021-
Map releaseMar 31, 2021-
UpdateJul 10, 2024-
Current statusJul 10, 2024Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.015
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.015
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6ye4
  • Surface level: 0.015
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_10789.png

Downloads & links

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Map

FileDownload / File: emd_10789.map.gz / Format: CCP4 / Size: 184 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 364 pix.
= 301.028 Å		0.83 Å/pix.
x 364 pix.
= 301.028 Å		0.83 Å/pix.
x 364 pix.
= 301.028 Å

Surface

Projections

Slices (1/3)

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.827 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.012 / Movie #1: 0.015
Minimum - Maximum-0.07147077 - 0.11590917
Average (Standard dev.)0.00010691346 (±0.0022554384)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions364364364
Spacing364364364
CellA=B=C: 301.028 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.8270.8270.827
M x/y/z364364364
origin x/y/z0.0000.0000.000
length x/y/z301.028301.028301.028
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS364364364
D min/max/mean-0.0710.1160.000

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Supplemental data

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Mask #1

Fileemd_10789_msk_1.map
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Additional map: #1

Fileemd_10789_additional_1.map
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Half map: #1

Fileemd_10789_half_map_1.map
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Half map: #2

Fileemd_10789_half_map_2.map
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Sample components

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Entire : Homopentamer of ExbB

EntireName: Homopentamer of ExbB
Components
  • Complex: Homopentamer of ExbB
    • Protein or peptide: Biopolymer transport protein ExbB
  • Ligand: (1S)-2-{[{[(2R)-2,3-DIHYDROXYPROPYL]OXY}(HYDROXY)PHOSPHORYL]OXY}-1-[(PALMITOYLOXY)METHYL]ETHYL STEARATE

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Supramolecule #1: Homopentamer of ExbB

SupramoleculeName: Homopentamer of ExbB / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: the expressed sequence corresponds to the mature sequence after signal peptide cleavage.
Source (natural)Organism: Serratia marcescens (bacteria) / Strain: Db11
Molecular weightTheoretical: 174 KDa

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Macromolecule #1: Biopolymer transport protein ExbB

MacromoleculeName: Biopolymer transport protein ExbB / type: protein_or_peptide / ID: 1 / Details: a 6-histidine tag is present at the C-terminus / Number of copies: 5 / Enantiomer: LEVO
Source (natural)Organism: Serratia marcescens (bacteria)
Molecular weightTheoretical: 30.413621 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: APAANPAVTE SVAPTTAPAP AAAAPESITP VNPAPTIQPP ETRGMDLSIW GMYQHADAVV KAVMIGLVLA SIVTWTILFA KGSELLRAK RRLRREQLAL AEARSLDEAS ELAQNFSPES VSAVLLNDAQ NELELSAESN DNNGIKERTG FRLERRVAAY S RNMGRGNG ...String:
APAANPAVTE SVAPTTAPAP AAAAPESITP VNPAPTIQPP ETRGMDLSIW GMYQHADAVV KAVMIGLVLA SIVTWTILFA KGSELLRAK RRLRREQLAL AEARSLDEAS ELAQNFSPES VSAVLLNDAQ NELELSAESN DNNGIKERTG FRLERRVAAY S RNMGRGNG FLATIGAISP FVGLFGTVWG IMNSFIGIAH SQTTNLAVVA PGIAEALLAT AMGLVAAIPA VVIYNIFARV IS GHRAQVG DVAAQVLLLQ GRDLDLAATA EAKRSQHAHQ LRAGHHHHHH

UniProtKB: Biopolymer transport protein ExbB

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Macromolecule #2: (1S)-2-{[{[(2R)-2,3-DIHYDROXYPROPYL]OXY}(HYDROXY)PHOSPHORYL]OXY}-...

MacromoleculeName: (1S)-2-{[{[(2R)-2,3-DIHYDROXYPROPYL]OXY}(HYDROXY)PHOSPHORYL]OXY}-1-[(PALMITOYLOXY)METHYL]ETHYL STEARATE
type: ligand / ID: 2 / Number of copies: 5 / Formula: PGT
Molecular weightTheoretical: 751.023 Da
Chemical component information

ChemComp-PGT:
(1S)-2-{[{[(2R)-2,3-DIHYDROXYPROPYL]OXY}(HYDROXY)PHOSPHORYL]OXY}-1-[(PALMITOYLOXY)METHYL]ETHYL STEARATE / phospholipid*YM

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.1 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
20.0 mMtris-HCl
100.0 mMNaClSodium chloride
0.0015 % w/vLMNGlauryl maltose neopentyl glycol

Details: 20mM Tris-HCl pH 8,0 100mM NaCl 0,0015% LMNG
GridModel: Quantifoil R2/4 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 277 K / Instrument: LEICA EM CPC
Detailsthe sample was monodisperse as evidenced by gel filtration column

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Electron microscopy

MicroscopeFEI TITAN KRIOS
TemperatureMin: 83.0 K / Max: 93.0 K
Specialist opticsEnergy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Dimensions - Width: 3710 pixel / Digitization - Dimensions - Height: 3838 pixel / Digitization - Frames/image: 1-56 / Number grids imaged: 1 / Number real images: 3122 / Average exposure time: 7.0 sec. / Average electron dose: 55.95 e/Å2
Details: frames were weighted according to electron dose and particle movement during Relion bayesian polishing procedure.
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 165000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Detailsimages were processed with Motioncor2
Particle selectionNumber selected: 1291382
Startup modelType of model: NONE / Details: used ab initio model on a subset of particles
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C5 (5 fold cyclic) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0) / Number images used: 157111
Initial angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: RELION (ver. 3.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final 3D classificationNumber classes: 4 / Software - Name: RELION (ver. 3.0)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

5sv0


Chain - Chain ID: A / Chain - Residue range: 10-234 / Chain - Source name: PDB / Chain - Initial model type: experimental model
Detailsthe 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.
RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Overall B value: 55.3 / Target criteria: Correlation coefficient
Output model

PDB-6ye4:
Structure of ExbB pentamer from Serratia marcescens by single particle cryo electron microscopy

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