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- EMDB-6552: Cryo-EM structure of the magnesium channel CorA in the magnesium-... -

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Basic information

Entry
Database: EMDB / ID: EMD-6552
TitleCryo-EM structure of the magnesium channel CorA in the magnesium-free asymmetric open state I
Map dataReconstruction of Thermotoga maritima CorA in the absence of magnesium resulting in at least two conformations; here state I
Sample
  • Sample: CorA from Thermotoga maritima in the absence of magnesium, state I
  • Protein or peptide: CorA
Keywordsmembrane protein / ion channel / magnesium channel / pentameric complex / symmetry vs. asymmetry / conformational change / direct electron detector / K2 / single-particle cryo-electron microscopy
Function / homology
Function and homology information


magnesium ion transmembrane transport / cobalt ion transport / cobalt ion transmembrane transporter activity / magnesium ion transmembrane transporter activity / cobalt ion binding / protein homooligomerization / magnesium ion binding / identical protein binding / plasma membrane
Similarity search - Function
Magnesium/cobalt transport protein CorA / Magnesium/cobalt transport protein CorA / Mg2+ transporter protein, CorA-like/Zinc transport protein ZntB / CorA, cytoplasmic domain / CorA, transmembrane region / CorA-like Mg2+ transporter protein
Similarity search - Domain/homology
Cobalt/magnesium transport protein CorA
Similarity search - Component
Biological speciesThermotoga maritima (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 7.06 Å
AuthorsMatthies D / Dalmas O / Borgnia MJ / Dominik PK / Merk A / Rao P / Reddy BG / Islam S / Bartesaghi A / Perozo E / Subramaniam S
CitationJournal: Cell / Year: 2016
Title: Cryo-EM Structures of the Magnesium Channel CorA Reveal Symmetry Break upon Gating.
Authors: Doreen Matthies / Olivier Dalmas / Mario J Borgnia / Pawel K Dominik / Alan Merk / Prashant Rao / Bharat G Reddy / Shahidul Islam / Alberto Bartesaghi / Eduardo Perozo / Sriram Subramaniam /
Abstract: CorA, the major Mg(2+) uptake system in prokaryotes, is gated by intracellular Mg(2+) (KD ∼ 1-2 mM). X-ray crystallographic studies of CorA show similar conformations under Mg(2+)-bound and Mg(2+)- ...CorA, the major Mg(2+) uptake system in prokaryotes, is gated by intracellular Mg(2+) (KD ∼ 1-2 mM). X-ray crystallographic studies of CorA show similar conformations under Mg(2+)-bound and Mg(2+)-free conditions, but EPR spectroscopic studies reveal large Mg(2+)-driven quaternary conformational changes. Here, we determined cryo-EM structures of CorA in the Mg(2+)-bound closed conformation and in two open Mg(2+)-free states at resolutions of 3.8, 7.1, and 7.1 Å, respectively. In the absence of bound Mg(2+), four of the five subunits are displaced to variable extents (∼ 10-25 Å) by hinge-like motions as large as ∼ 35° at the stalk helix. The transition between a single 5-fold symmetric closed state and an ensemble of low Mg(2+), open, asymmetric conformational states is, thus, the key structural signature of CorA gating. This mechanism is likely to apply to other structurally similar divalent ion channels.
History
DepositionNov 30, 2015-
Header (metadata) releaseJan 13, 2016-
Map releaseFeb 17, 2016-
UpdateMar 2, 2016-
Current statusMar 2, 2016Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.03
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.03
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-3jcg
  • Surface level: 0.03
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_6552.png

emd_6552_1.png

Downloads & links

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Map

FileDownload / File: emd_6552.map.gz / Format: CCP4 / Size: 62.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationReconstruction of Thermotoga maritima CorA in the absence of magnesium resulting in at least two conformations; here state I
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.35 Å/pix.
x 256 pix.
= 346.112 Å		1.35 Å/pix.
x 256 pix.
= 346.112 Å		1.35 Å/pix.
x 256 pix.
= 346.112 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.352 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.03 / Movie #1: 0.03
Minimum - Maximum-0.02053478 - 0.0724296
Average (Standard dev.)0.00042226 (±0.00328655)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 346.112 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.3521.3521.352
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z346.112346.112346.112
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.0210.0720.000

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Supplemental data

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Sample components

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Entire : CorA from Thermotoga maritima in the absence of magnesium, state I

EntireName: CorA from Thermotoga maritima in the absence of magnesium, state I
Components
  • Sample: CorA from Thermotoga maritima in the absence of magnesium, state I
  • Protein or peptide: CorA

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Supramolecule #1000: CorA from Thermotoga maritima in the absence of magnesium, state I

SupramoleculeName: CorA from Thermotoga maritima in the absence of magnesium, state I
type: sample / ID: 1000 / Details: Detergent-solubilized, purified protein / Oligomeric state: One homopentamer of CorA / Number unique components: 1
Molecular weightTheoretical: 200 KDa

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Macromolecule #1: CorA

MacromoleculeName: CorA / type: protein_or_peptide / ID: 1 / Name.synonym: Magnesium Channel CorA / Number of copies: 5 / Oligomeric state: Pentamer / Recombinant expression: Yes
Source (natural)Organism: Thermotoga maritima (bacteria) / Location in cell: Inner Membrane
Molecular weightTheoretical: 200 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli) / Recombinant strain: BL21(DE3) PlysS / Recombinant plasmid: Cora-Pet15b
SequenceUniProtKB: Cobalt/magnesium transport protein CorA / InterPro: Magnesium/cobalt transport protein CorA

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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

Concentration3.7 mg/mL
BufferpH: 7.3 / Details: 50 mM HEPES, 150 mM NaCl, 1 mM EDTA, 0.5 mM DDM
GridDetails: 300 mesh Cu R1.2/1.3 holey carbon grids from Quantifoil, plasma-cleaned
VitrificationCryogen name: ETHANE / Chamber humidity: 86 % / Chamber temperature: 90 K / Instrument: LEICA EM GP
Method: Grids were blotted at 4 degrees Celsius for 7 seconds after a 10-second pre-blotting period, then plunge-frozen in liquid ethane.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
TemperatureMin: 79.6 K / Max: 79.8 K / Average: 79.7 K
Alignment procedureLegacy - Astigmatism: Objective lens astigmatism was corrected at 105,000 times magnification.
Legacy - Electron beam tilt params: 5
Specialist opticsEnergy filter - Name: Gatan, Inc. / Energy filter - Lower energy threshold: 0.0 eV / Energy filter - Upper energy threshold: 20.0 eV
DetailsParallel beam illumination
DateOct 15, 2014
Image recordingCategory: CCD / Film or detector model: GATAN K2 QUANTUM (4k x 4k) / Number real images: 2498 / Average electron dose: 40 e/Å2
Details: Every image is the average of 38 frames recorded by the direct electron detector. The total exposure time was 15.2 seconds, and intermediate frames were recorded every 0.4 seconds.
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 105000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.7 µm / Nominal defocus min: 0.89 µm / Nominal magnification: 105000
Sample stageSpecimen holder: Liquid nitrogen-cooled / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

DetailsParticles were selected using an automatic selection program. 3D classification, 3D refinement, and post-processing were done using RELION 1.3.
CTF correctionDetails: CTF parameters obtained from whole micrograph
Final reconstructionResolution.type: BY AUTHOR / Resolution: 7.06 Å / Resolution method: OTHER / Software - Name: RELION
Details: Final maps were calculated from two merged datasets.
Number images used: 26271
FSC plot (resolution estimation)

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