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- PDB-7ua1: Structure of PKA phosphorylated human RyR2-R2474S in the closed s... -

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Basic information

Entry
Database: PDB / ID: 7ua1
TitleStructure of PKA phosphorylated human RyR2-R2474S in the closed state in the presence of ARM210
Components
  • Peptidyl-prolyl cis-trans isomerase FKBP1B
  • Ryanodine receptor 2
KeywordsMEMBRANE PROTEIN / calcium channel
Function / homology
Function and homology information


junctional sarcoplasmic reticulum membrane / 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 / left ventricular cardiac muscle tissue morphogenesis / regulation of AV node cell action potential / calcium-induced calcium release activity ...junctional sarcoplasmic reticulum membrane / 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 / left ventricular cardiac muscle tissue morphogenesis / regulation of AV node cell action potential / calcium-induced calcium release activity / sarcoplasmic reticulum calcium ion transport / cell communication by electrical coupling involved in cardiac conduction / regulation of ventricular cardiac muscle cell action potential / ventricular cardiac muscle cell action potential / positive regulation of sequestering of calcium ion / cyclic nucleotide binding / negative regulation of calcium-mediated signaling / embryonic heart tube morphogenesis / cardiac muscle hypertrophy / negative regulation of insulin secretion involved in cellular response to glucose stimulus / regulation of cardiac muscle contraction by calcium ion signaling / negative regulation of release of sequestered calcium ion into cytosol / neuronal action potential propagation / insulin secretion involved in cellular response to glucose stimulus / ryanodine-sensitive calcium-release channel activity / response to caffeine / calcium ion transport into cytosol / response to muscle activity / release of sequestered calcium ion into cytosol by sarcoplasmic reticulum / response to redox state / protein maturation by protein folding / 'de novo' protein folding / negative regulation of heart rate / positive regulation of heart rate / FK506 binding / positive regulation of axon regeneration / protein kinase A regulatory subunit binding / cellular response to caffeine / protein kinase A catalytic subunit binding / channel regulator activity / positive regulation of the force of heart contraction / intracellularly gated calcium channel activity / detection of calcium ion / regulation of cardiac muscle contraction / smooth muscle contraction / response to vitamin E / smooth endoplasmic reticulum / regulation of cardiac muscle contraction by regulation of the release of sequestered calcium ion / calcium channel inhibitor activity / cardiac muscle contraction / striated muscle contraction / T cell proliferation / regulation of release of sequestered calcium ion into cytosol by sarcoplasmic reticulum / Ion homeostasis / release of sequestered calcium ion into cytosol / regulation of cytosolic calcium ion concentration / calcium channel complex / cellular response to epinephrine stimulus / response to muscle stretch / sarcoplasmic reticulum membrane / regulation of heart rate / sarcoplasmic reticulum / establishment of localization in cell / peptidylprolyl isomerase / peptidyl-prolyl cis-trans isomerase activity / calcium-mediated signaling / calcium channel activity / response to hydrogen peroxide / sarcolemma / Stimuli-sensing channels / Z disc / intracellular calcium ion homeostasis / calcium ion transport / positive regulation of cytosolic calcium ion concentration / protein refolding / transmembrane transporter binding / calmodulin binding / response to hypoxia / signaling receptor binding / calcium ion binding / enzyme binding / protein-containing complex / identical protein binding / membrane / plasma membrane / cytosol / cytoplasm
Similarity search - Function
Ryanodine receptor, SPRY domain 2 / : / Ryanodine receptor junctional solenoid repeat / Ryanodine Receptor TM 4-6 / Ryanodine receptor / Ryanodine receptor, SPRY domain 1 / Ryanodine receptor, SPRY domain 3 / Ryanodine Receptor TM 4-6 / Ryanodine receptor Ryr / RyR domain ...Ryanodine receptor, SPRY domain 2 / : / Ryanodine receptor junctional solenoid repeat / Ryanodine Receptor TM 4-6 / Ryanodine receptor / Ryanodine receptor, SPRY domain 1 / Ryanodine receptor, SPRY domain 3 / Ryanodine Receptor TM 4-6 / Ryanodine receptor Ryr / RyR domain / 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 / SPRY domain / B30.2/SPRY domain / B30.2/SPRY domain profile. / B30.2/SPRY domain superfamily / Domain in SPla and the RYanodine Receptor. / SPRY domain / FKBP-type peptidyl-prolyl cis-trans isomerase domain profile. / FKBP-type peptidyl-prolyl cis-trans isomerase domain / FKBP-type peptidyl-prolyl cis-trans isomerase / Peptidyl-prolyl cis-trans isomerase domain superfamily / EF-hand domain pair / EF-hand domain / Ion transport domain / Ion transport protein / EF-hand domain pair / Concanavalin A-like lectin/glucanase domain superfamily
Similarity search - Domain/homology
ADENOSINE-5'-TRIPHOSPHATE / Chem-KVR / Peptidyl-prolyl cis-trans isomerase FKBP1B / Ryanodine receptor 2
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.99 Å
AuthorsMiotto, M.C. / Marks, A.R.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)R01HL145473 United States
CitationJournal: Sci Adv / Year: 2022
Title: Structural analyses of human ryanodine receptor type 2 channels reveal the mechanisms for sudden cardiac death and treatment.
Authors: Marco C Miotto / Gunnar Weninger / Haikel Dridi / Qi Yuan / Yang Liu / Anetta Wronska / Zephan Melville / Leah Sittenfeld / Steven Reiken / Andrew R Marks /
Abstract: Ryanodine receptor type 2 (RyR2) mutations have been linked to an inherited form of exercise-induced sudden cardiac death called catecholaminergic polymorphic ventricular tachycardia (CPVT). CPVT ...Ryanodine receptor type 2 (RyR2) mutations have been linked to an inherited form of exercise-induced sudden cardiac death called catecholaminergic polymorphic ventricular tachycardia (CPVT). CPVT results from stress-induced sarcoplasmic reticular Ca leak via the mutant RyR2 channels during diastole. We present atomic models of human wild-type (WT) RyR2 and the CPVT mutant RyR2-R2474S determined by cryo-electron microscopy with overall resolutions in the range of 2.6 to 3.6 Å, and reaching local resolutions of 2.25 Å, unprecedented for RyR2 channels. Under nonactivating conditions, the RyR2-R2474S channel is in a "primed" state between the closed and open states of WT RyR2, rendering it more sensitive to activation that results in stress-induced Ca leak. The Rycal drug ARM210 binds to RyR2-R2474S, reverting the primed state toward the closed state. Together, these studies provide a mechanism for CPVT and for the therapeutic actions of ARM210.
History
DepositionMar 11, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Aug 3, 2022Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Ryanodine receptor 2
E: Peptidyl-prolyl cis-trans isomerase FKBP1B
F: Peptidyl-prolyl cis-trans isomerase FKBP1B
G: Peptidyl-prolyl cis-trans isomerase FKBP1B
H: Peptidyl-prolyl cis-trans isomerase FKBP1B
B: Ryanodine receptor 2
C: Ryanodine receptor 2
D: Ryanodine receptor 2
hetero molecules


Theoretical massNumber of molelcules
Total (without water)2,313,69524
Polymers2,308,0588
Non-polymers5,63716
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Ryanodine receptor 2 / RYR-2 / RyR2 / hRYR-2 / Cardiac muscle ryanodine receptor / Cardiac muscle ryanodine receptor- ...RYR-2 / RyR2 / hRYR-2 / Cardiac muscle ryanodine receptor / Cardiac muscle ryanodine receptor-calcium release channel / Type 2 ryanodine receptor


Mass: 565216.000 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: RYR2 / Cell line (production host): HEK293 / Production host: Homo sapiens (human) / References: UniProt: Q92736
#2: Protein
Peptidyl-prolyl cis-trans isomerase FKBP1B / PPIase FKBP1B / 12.6 kDa FK506-binding protein / FKBP-12.6 / FK506-binding protein 1B / FKBP-1B / ...PPIase FKBP1B / 12.6 kDa FK506-binding protein / FKBP-12.6 / FK506-binding protein 1B / FKBP-1B / Immunophilin FKBP12.6 / Rotamase / h-FKBP-12


Mass: 11798.501 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: FKBP1B, FKBP12.6, FKBP1L, FKBP9, OTK4 / Production host: Escherichia coli (E. coli) / References: UniProt: P68106, peptidylprolyl isomerase
#3: Chemical
ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: Zn / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical
ChemComp-ATP / ADENOSINE-5'-TRIPHOSPHATE


Mass: 507.181 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: C10H16N5O13P3 / Feature type: SUBJECT OF INVESTIGATION / Comment: ATP, energy-carrying molecule*YM
#5: Chemical
ChemComp-KVR / 4-[(7-methoxy-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)methyl]benzoic acid


Mass: 329.413 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C18H19NO3S
Has ligand of interestY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

Component
IDNameTypeEntity IDParent-IDSource
1Complex of RyR2-R2474S and Calstabin-2 in the presence of ARM210COMPLEX#1-#20RECOMBINANT
2Ryanodine receptor 2COMPLEX#11RECOMBINANT
3Peptidyl-prolyl cis-trans isomerase FKBP1BCOMPLEX#21RECOMBINANT
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
12Homo sapiens (human)9606
23Homo sapiens (human)9606
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-IDCell
12Homo sapiens (human)9606HEK293
23Escherichia coli (E. coli)562
Buffer solutionpH: 7.4
Details: Xanthine was made fresh to avoid aggregation. Xanthine stock solution was 10 mM in NaOH 0.5 N.
Buffer component
IDConc.NameFormulaBuffer-ID
1230 mMNaClNaCl1
210 mMHEPESHEPES1
30.4 %CHAPSCHAPS1
41 mMEGTAEGTA1
50.5 mMTCEPTCEP1
60.001 %DOPCDOPC1
710 mMATPATP1
80.2 mMcAMPcAMP1
90.65 mMCaCl2CaCl21
100.5 mMXanthineXanthine1
110.5 mM4-[(7-methoxy-3,5-dihydro-2H-1,4-benzothiazepin-4-yl)methyl]benzoic acidARM2101
SpecimenConc.: 2.2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 1200 nm / Nominal defocus min: 400 nm
Image recordingElectron dose: 58 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

EM softwareName: cryoSPARC / Category: CTF correction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.99 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 113172 / Symmetry type: POINT
Atomic model buildingPDB-ID: 7U9X

7u9x

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