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- EMDB-9641: Calcium release-activated calcium channel protein 1, P288L mutant -

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Basic information

Entry
Database: EMDB / ID: EMD-9641
TitleCalcium release-activated calcium channel protein 1, P288L mutant
Map data
Sample
  • Organelle or cellular component: Calcium release-activated calcium channel protein 1
Biological speciesDrosophila (fruit flies)
Methodsingle particle reconstruction / cryo EM / Resolution: 5.6 Å
AuthorsLiu X / Wu G / Yang X / Shen Y
CitationJournal: PLoS Biol / Year: 2019
Title: Molecular understanding of calcium permeation through the open Orai channel.
Authors: Xiaofen Liu / Guangyan Wu / Yi Yu / Xiaozhe Chen / Renci Ji / Jing Lu / Xin Li / Xing Zhang / Xue Yang / Yuequan Shen /
Abstract: The Orai channel is characterized by voltage independence, low conductance, and high Ca2+ selectivity and plays an important role in Ca2+ influx through the plasma membrane (PM). How the channel is ...The Orai channel is characterized by voltage independence, low conductance, and high Ca2+ selectivity and plays an important role in Ca2+ influx through the plasma membrane (PM). How the channel is activated and promotes Ca2+ permeation is not well understood. Here, we report the crystal structure and cryo-electron microscopy (cryo-EM) reconstruction of a Drosophila melanogaster Orai (dOrai) mutant (P288L) channel that is constitutively active according to electrophysiology. The open state of the Orai channel showed a hexameric assembly in which 6 transmembrane 1 (TM1) helices in the center form the ion-conducting pore, and 6 TM4 helices in the periphery form extended long helices. Orai channel activation requires conformational transduction from TM4 to TM1 and eventually causes the basic section of TM1 to twist outward. The wider pore on the cytosolic side aggregates anions to increase the potential gradient across the membrane and thus facilitate Ca2+ permeation. The open-state structure of the Orai channel offers insights into channel assembly, channel activation, and Ca2+ permeation.
History
DepositionSep 1, 2018-
Header (metadata) releaseMar 20, 2019-
Map releaseMar 20, 2019-
UpdateApr 1, 2020-
Current statusApr 1, 2020Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.00306
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.00306
  • Imaged by UCSF Chimera
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Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

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Map

FileDownload / File: emd_9641.map.gz / Format: CCP4 / Size: 40.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.014 Å
Density
Contour LevelBy AUTHOR: 0.01 / Movie #1: 0.00306
Minimum - Maximum-0.012362895 - 0.040860157
Average (Standard dev.)0.0001223818 (±0.001473981)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions220220220
Spacing220220220
CellA=B=C: 223.08002 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0141.0141.014
M x/y/z220220220
origin x/y/z0.0000.0000.000
length x/y/z223.080223.080223.080
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ256256256
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS220220220
D min/max/mean-0.0120.0410.000

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Supplemental data

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Sample components

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Entire : Calcium release-activated calcium channel protein 1

EntireName: Calcium release-activated calcium channel protein 1
Components
  • Organelle or cellular component: Calcium release-activated calcium channel protein 1

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Supramolecule #1: Calcium release-activated calcium channel protein 1

SupramoleculeName: Calcium release-activated calcium channel protein 1 / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Drosophila (fruit flies)
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: HEK293S-GnTi- / Recombinant plasmid: pEG-BacMam

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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

BufferpH: 8
GridMaterial: COPPER
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELDBright-field microscopy
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Average electron dose: 49.8 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Details: Generated by a pdb model from crystal diffraction data
Initial angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 2.1)
Final 3D classificationNumber classes: 6 / Avg.num./class: 20000 / Software - Name: RELION
Final angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 2.1)
Final reconstructionApplied symmetry - Point group: C3 (3 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 5.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 2.1) / Number images used: 22442
FSC plot (resolution estimation)

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