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Yorodumi- PDB-8rrs: Structure of mouse RyR2 solubilised in detergent in open state in... -
+Open data
-Basic information
Entry | Database: PDB / ID: 8rrs | ||||||||||||
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Title | Structure of mouse RyR2 solubilised in detergent in open state in complex with Ca2+, ATP, caffeine and Nb9657. | ||||||||||||
Components |
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Keywords | TRANSPORT PROTEIN / Ion channel / Ca2+ / tetramer | ||||||||||||
Function / homology | Function and homology information manganese ion transmembrane transport / establishment of protein localization to endoplasmic reticulum / type B pancreatic cell apoptotic process / Purkinje myocyte to ventricular cardiac muscle cell signaling / suramin binding / regulation of SA node cell action potential / regulation of atrial cardiac muscle cell action potential / sarcoplasmic reticulum calcium ion transport / left ventricular cardiac muscle tissue morphogenesis / organic cyclic compound binding ...manganese ion transmembrane transport / establishment of protein localization to endoplasmic reticulum / type B pancreatic cell apoptotic process / Purkinje myocyte to ventricular cardiac muscle cell signaling / suramin binding / regulation of SA node cell action potential / regulation of atrial cardiac muscle cell action potential / sarcoplasmic reticulum calcium ion transport / left ventricular cardiac muscle tissue morphogenesis / organic cyclic compound binding / regulation of AV node cell action potential / calcium-induced calcium release activity / Stimuli-sensing channels / Ion homeostasis / regulation of ventricular cardiac muscle cell action potential / positive regulation of sequestering of calcium ion / ventricular cardiac muscle cell action potential / cardiac muscle hypertrophy / embryonic heart tube morphogenesis / ryanodine-sensitive calcium-release channel activity / response to muscle activity / release of sequestered calcium ion into cytosol by sarcoplasmic reticulum / calcium ion transport into cytosol / response to caffeine / response to redox state / A band / calcium ion transmembrane import into cytosol / positive regulation of heart rate / negative regulation of cytosolic calcium ion concentration / cellular response to caffeine / protein kinase A regulatory subunit binding / protein kinase A catalytic subunit binding / extrinsic component of cytoplasmic side of plasma membrane / positive regulation of the force of heart contraction / intracellularly gated calcium channel activity / response to magnesium ion / detection of calcium ion / smooth endoplasmic reticulum / regulation of cardiac muscle contraction by regulation of the release of sequestered calcium ion / striated muscle contraction / regulation of cytosolic calcium ion concentration / sarcoplasmic reticulum membrane / calcium channel complex / cellular response to epinephrine stimulus / response to muscle stretch / sarcomere / regulation of heart rate / sarcoplasmic reticulum / establishment of localization in cell / calcium ion transmembrane transport / calcium-mediated signaling / calcium channel activity / sarcolemma / Z disc / intracellular calcium ion homeostasis / response to calcium ion / monoatomic ion transmembrane transport / calcium ion transport / nuclear envelope / scaffold protein binding / response to hypoxia / calmodulin binding / calcium ion binding / protein kinase binding / enzyme binding / protein-containing complex / identical protein binding / membrane Similarity search - Function | ||||||||||||
Biological species | Vicugna pacos (alpaca) Mus musculus (house mouse) | ||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.4 Å | ||||||||||||
Authors | Li, C. / Efremov, R.G. | ||||||||||||
Funding support | Belgium, European Union, 3items
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Citation | Journal: J Biol Chem / Year: 2024 Title: Rapid small-scale nanobody-assisted purification of ryanodine receptors for cryo-EM. Authors: Chenyao Li / Katrien Willegems / Tomasz Uchański / Els Pardon / Jan Steyaert / Rouslan G Efremov / Abstract: Ryanodine receptors (RyRs) are large Ca release channels residing in the endoplasmic or sarcoplasmic reticulum membrane. Three isoforms of RyRs have been identified in mammals, the disfunction of ...Ryanodine receptors (RyRs) are large Ca release channels residing in the endoplasmic or sarcoplasmic reticulum membrane. Three isoforms of RyRs have been identified in mammals, the disfunction of which has been associated with a series of life-threatening diseases. The need for large amounts of native tissue or eukaryotic cell cultures limits advances in structural studies of RyRs. Here, we report a method that utilizes nanobodies to purify RyRs from only 5 mg of total protein. The purification process, from isolated membranes to cryo-EM grade protein, is achieved within 4 h on the bench, yielding protein usable for cryo-EM analysis. This is demonstrated by solving the structures of rabbit RyR1, solubilized in detergent, reconstituted into lipid nanodiscs or liposomes, and bovine RyR2 reconstituted in nanodisc, and mouse RyR2 in detergent. The reported method facilitates structural studies of RyRs directed toward drug development and is useful in cases where the amount of starting material is limited. | ||||||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8rrs.cif.gz | 3.3 MB | Display | PDBx/mmCIF format |
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PDB format | pdb8rrs.ent.gz | Display | PDB format | |
PDBx/mmJSON format | 8rrs.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8rrs_validation.pdf.gz | 1.5 MB | Display | wwPDB validaton report |
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Full document | 8rrs_full_validation.pdf.gz | 1.8 MB | Display | |
Data in XML | 8rrs_validation.xml.gz | 401.7 KB | Display | |
Data in CIF | 8rrs_validation.cif.gz | 632.3 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/rr/8rrs ftp://data.pdbj.org/pub/pdb/validation_reports/rr/8rrs | HTTPS FTP |
-Related structure data
Related structure data | 19463MC 8rrtC 8rruC 8rrvC 8rrwC 8rrxC 8rs0C 19509 19511 M: map data used to model this data C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-Protein / Antibody , 2 types, 8 molecules ACEFBDGI
#1: Protein | Mass: 565536.000 Da / Num. of mol.: 4 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse) / References: UniProt: E9Q401 #2: Antibody | Mass: 15125.495 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Vicugna pacos (alpaca) / Production host: Escherichia coli (E. coli) |
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-Non-polymers , 4 types, 16 molecules
#3: Chemical | ChemComp-ATP / #4: Chemical | ChemComp-CFF / #5: Chemical | ChemComp-ZN / #6: Chemical | ChemComp-CA / |
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-Details
Has ligand of interest | Y |
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Has protein modification | Y |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component |
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Source (natural) |
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Source (recombinant) | Organism: Escherichia coli (E. coli) | ||||||||||||||||||||||||
Buffer solution | pH: 7.4 | ||||||||||||||||||||||||
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||
Vitrification | Cryogen name: ETHANE |
-Electron microscopy imaging
Microscopy | Model: JEOL CRYO ARM 300 |
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Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: SPOT SCAN |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2500 nm / Nominal defocus min: 1500 nm |
Image recording | Electron dose: 60 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
-Processing
EM software |
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Image processing | Details: The movies were selected after MotionCor correction and CTF refinement with the parameters: total drift less than 30 angstrom and resolution better than 5 angstrom | ||||||||||||||||||||||||||||||
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 791664 Details: Particles were picked by template picking in Cryosparc | ||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 76852 / Symmetry type: POINT |