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- EMDB-6369: Structure of full-length IP3R1 channel in the apo-state determine... -

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Basic information

Entry
Database: EMDB / ID: EMD-6369
TitleStructure of full-length IP3R1 channel in the apo-state determined by single particle cryo-EM
Map dataReconstruction of IP3R1/Calcium release channel from the rat cerebellum
Sample
  • Sample: Inositol 1,4,5-trisphosphate receptor, type 1
  • Protein or peptide: Inositol 1,4,5-trisphosphate receptor
Keywordsinositol 1 / 4 / 5-trisphosphate receptor / calcium release channel / single-particle cryo-EM / calcium signaling / 3D reconstruction
Function / homology
Function and homology information


Effects of PIP2 hydrolysis / Antigen activates B Cell Receptor (BCR) leading to generation of second messengers / inositol 1,4,5-trisphosphate receptor activity involved in regulation of postsynaptic cytosolic calcium levels / release of sequestered calcium ion into cytosol by endoplasmic reticulum / smooth endoplasmic reticulum membrane / cGMP effects / Elevation of cytosolic Ca2+ levels / platelet dense tubular network / calcineurin complex / platelet dense granule membrane ...Effects of PIP2 hydrolysis / Antigen activates B Cell Receptor (BCR) leading to generation of second messengers / inositol 1,4,5-trisphosphate receptor activity involved in regulation of postsynaptic cytosolic calcium levels / release of sequestered calcium ion into cytosol by endoplasmic reticulum / smooth endoplasmic reticulum membrane / cGMP effects / Elevation of cytosolic Ca2+ levels / platelet dense tubular network / calcineurin complex / platelet dense granule membrane / epithelial fluid transport / inositol 1,4,5-trisphosphate-gated calcium channel activity / phospholipase C-activating G protein-coupled acetylcholine receptor signaling pathway / calcium import into the mitochondrion / voluntary musculoskeletal movement / positive regulation of hepatocyte proliferation / inositol 1,4,5 trisphosphate binding / negative regulation of calcium-mediated signaling / positive regulation of calcium ion transport / endoplasmic reticulum calcium ion homeostasis / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / nuclear inner membrane / Ion homeostasis / transport vesicle membrane / dendrite development / intracellularly gated calcium channel activity / ligand-gated ion channel signaling pathway / intrinsic apoptotic signaling pathway in response to endoplasmic reticulum stress / regulation of cytosolic calcium ion concentration / single fertilization / calcium channel inhibitor activity / release of sequestered calcium ion into cytosol / liver regeneration / phosphatidylinositol binding / secretory granule membrane / synaptic membrane / cellular response to cAMP / sarcoplasmic reticulum / post-embryonic development / positive regulation of neuron projection development / positive regulation of insulin secretion / GABA-ergic synapse / Schaffer collateral - CA1 synapse / cell morphogenesis / calcium ion transport / nuclear envelope / presynapse / positive regulation of cytosolic calcium ion concentration / protein phosphatase binding / protein homotetramerization / phospholipase C-activating G protein-coupled receptor signaling pathway / cellular response to hypoxia / transmembrane transporter binding / response to hypoxia / postsynapse / postsynaptic density / positive regulation of apoptotic process / protein domain specific binding / neuronal cell body / calcium ion binding / synapse / dendrite / endoplasmic reticulum membrane / negative regulation of apoptotic process / protein-containing complex binding / nucleolus / perinuclear region of cytoplasm / endoplasmic reticulum / protein homodimerization activity / protein-containing complex / ATP binding / identical protein binding / membrane / plasma membrane / cytoplasm
Similarity search - Function
Inositol 1,4,5-trisphosphate receptor / RyR/IP3 receptor binding core, RIH domain superfamily / RyR/IP3R Homology associated domain / Inositol 1,4,5-trisphosphate/ryanodine receptor / RIH domain / RyR and IP3R Homology associated / Inositol 1,4,5-trisphosphate/ryanodine receptor / RIH domain / : / MIR motif ...Inositol 1,4,5-trisphosphate receptor / RyR/IP3 receptor binding core, RIH domain superfamily / RyR/IP3R Homology associated domain / Inositol 1,4,5-trisphosphate/ryanodine receptor / RIH domain / RyR and IP3R Homology associated / Inositol 1,4,5-trisphosphate/ryanodine receptor / RIH domain / : / MIR motif / MIR domain / MIR domain profile. / Domain in ryanodine and inositol trisphosphate receptors and protein O-mannosyltransferases / Mir domain superfamily / Ion transport domain / Ion transport protein / Armadillo-type fold
Similarity search - Domain/homology
Inositol 1,4,5-trisphosphate-gated calcium channel ITPR1
Similarity search - Component
Biological speciesRattus norvegicus (Norway rat)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.7 Å
AuthorsFan G / Baker ML / Wang Z / Baker MR / Sinyagovskiy PA / Chiu W / Ludtke SJ / Serysheva II
CitationJournal: Nature / Year: 2015
Title: Gating machinery of InsP3R channels revealed by electron cryomicroscopy.
Authors: Guizhen Fan / Matthew L Baker / Zhao Wang / Mariah R Baker / Pavel A Sinyagovskiy / Wah Chiu / Steven J Ludtke / Irina I Serysheva /
Abstract: Inositol-1,4,5-trisphosphate receptors (InsP3Rs) are ubiquitous ion channels responsible for cytosolic Ca(2+) signalling and essential for a broad array of cellular processes ranging from contraction ...Inositol-1,4,5-trisphosphate receptors (InsP3Rs) are ubiquitous ion channels responsible for cytosolic Ca(2+) signalling and essential for a broad array of cellular processes ranging from contraction to secretion, and from proliferation to cell death. Despite decades of research on InsP3Rs, a mechanistic understanding of their structure-function relationship is lacking. Here we present the first, to our knowledge, near-atomic (4.7 Å) resolution electron cryomicroscopy structure of the tetrameric mammalian type 1 InsP3R channel in its apo-state. At this resolution, we are able to trace unambiguously ∼85% of the protein backbone, allowing us to identify the structural elements involved in gating and modulation of this 1.3-megadalton channel. Although the central Ca(2+)-conduction pathway is similar to other ion channels, including the closely related ryanodine receptor, the cytosolic carboxy termini are uniquely arranged in a left-handed α-helical bundle, directly interacting with the amino-terminal domains of adjacent subunits. This configuration suggests a molecular mechanism for allosteric regulation of channel gating by intracellular signals.
History
DepositionJun 29, 2015-
Header (metadata) releaseAug 12, 2015-
Map releaseOct 7, 2015-
UpdateDec 9, 2015-
Current statusDec 9, 2015Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.7
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.7
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-3jav
  • Surface level: 0.7
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

400_6369.gif

Downloads & links

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Map

FileDownload / File: emd_6369.map.gz / Format: CCP4 / Size: 62.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationReconstruction of IP3R1/Calcium release channel from the rat cerebellum
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.62 Å/pix.
x 256 pix.
= 414.72 Å		1.62 Å/pix.
x 256 pix.
= 414.72 Å		1.62 Å/pix.
x 256 pix.
= 414.72 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.62 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.7 / Movie #1: 0.7
Minimum - Maximum-6.21526337 - 16.27171135
Average (Standard dev.)0.02987346 (±0.38540578)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 414.72 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.621.621.62
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z414.720414.720414.720
α/β/γ90.00090.00090.000
start NX/NY/NZ-8211244
NX/NY/NZ115138149
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-6.21516.2720.030

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Supplemental data

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Sample components

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Entire : Inositol 1,4,5-trisphosphate receptor, type 1

EntireName: Inositol 1,4,5-trisphosphate receptor, type 1
Components
  • Sample: Inositol 1,4,5-trisphosphate receptor, type 1
  • Protein or peptide: Inositol 1,4,5-trisphosphate receptor

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Supramolecule #1000: Inositol 1,4,5-trisphosphate receptor, type 1

SupramoleculeName: Inositol 1,4,5-trisphosphate receptor, type 1 / type: sample / ID: 1000
Details: The sample was purified from rat cerebellum after solubilization with detergent; only freshly purified protein was used for cryo-EM visualization.
Oligomeric state: tetramer / Number unique components: 1
Molecular weightTheoretical: 1.3 MDa

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Macromolecule #1: Inositol 1,4,5-trisphosphate receptor

MacromoleculeName: Inositol 1,4,5-trisphosphate receptor / type: protein_or_peptide / ID: 1 / Name.synonym: Calcium release channel / Details: detergent-solubilized protein / Number of copies: 4 / Oligomeric state: tetramer / Recombinant expression: No / Database: NCBI
Source (natural)Organism: Rattus norvegicus (Norway rat) / synonym: Rat / Tissue: cerebellum / Location in cell: endoplasmic reticulum membrane
Molecular weightExperimental: 300 KDa / Theoretical: 330 KDa
SequenceUniProtKB: Inositol 1,4,5-trisphosphate-gated calcium channel ITPR1

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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

Concentration0.4 mg/mL
BufferpH: 7.4
Details: 50 mM Tris-HCl, pH 7.4, 0.4% CHAPS, 150 mM NaCl, 1 mM DTT, 1 mM EGTA, 1 mM EDTA
GridDetails: 400 mesh copper grids with thin carbon support
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 120 K / Instrument: FEI VITROBOT MARK IV / Method: blot once for 3 seconds

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Electron microscopy

MicroscopeFEI POLARA 300
TemperatureMin: 95 K / Max: 102 K / Average: 100 K
Alignment procedureLegacy - Astigmatism: Objective lens astigmatism was corrected at 100,000 times magnification.
Specialist opticsEnergy filter - Name: FEI
DateJan 1, 2014
Image recordingCategory: CCD / Film or detector model: GATAN K2 (4k x 4k) / Digitization - Sampling interval: 0.81 µm / Number real images: 4160 / Average electron dose: 22 e/Å2
Details: Every image is the sum of 30 frames recorded with a direct electron detector.
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 30886 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.0 mm / Nominal defocus max: 3.5 µm / Nominal defocus min: 0.6 µm / Nominal magnification: 23000
Sample stageSpecimen holder model: GATAN LIQUID NITROGEN
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

DetailsParticle selection: EMAN2.1, CTF correction: CTFFIND3, initial model: EMAN2.1, refinement: RELION 1.3
CTF correctionDetails: CTFFIND3
Final reconstructionAlgorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 4.7 Å / Resolution method: OTHER / Software - Name: RELION_1.3, EMAN_2.1 / Details: The final map was generated from 96,106 particles. / Number images used: 96106

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