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- PDB-5l1d: Structure of rabbit RyR2 in complex with FKBP12.6 in a closed sta... -

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Basic information

Entry
Database: PDB / ID: 5l1d
TitleStructure of rabbit RyR2 in complex with FKBP12.6 in a closed state (conformation C1)
Components
  • Peptidyl-prolyl cis-trans isomerase FKBP1B
  • Ryanodine Receptor
KeywordsTRANSPORT PROTEIN/ISOMERASE / Calcium Release Channel / Ryanodine Receptor / TRANSPORT PROTEIN-ISOMERASE complex
Function / homology
Function and homology information


positive regulation of sequestering of calcium ion / cyclic nucleotide binding / negative regulation of release of sequestered calcium ion into cytosol / negative regulation of insulin secretion involved in cellular response to glucose stimulus / neuronal action potential propagation / insulin secretion involved in cellular response to glucose stimulus / cell communication by electrical coupling involved in cardiac conduction / response to redox state / protein maturation by protein folding / 'de novo' protein folding ...positive regulation of sequestering of calcium ion / cyclic nucleotide binding / negative regulation of release of sequestered calcium ion into cytosol / negative regulation of insulin secretion involved in cellular response to glucose stimulus / neuronal action potential propagation / insulin secretion involved in cellular response to glucose stimulus / cell communication by electrical coupling involved in cardiac conduction / response to redox state / protein maturation by protein folding / 'de novo' protein folding / negative regulation of heart rate / negative regulation of phosphoprotein phosphatase activity / FK506 binding / positive regulation of axon regeneration / : / smooth muscle contraction / negative regulation of ryanodine-sensitive calcium-release channel activity / response to vitamin E / calcium channel inhibitor activity / regulation of cardiac muscle contraction by regulation of the release of sequestered calcium ion / protein peptidyl-prolyl isomerization / T cell proliferation / regulation of release of sequestered calcium ion into cytosol by sarcoplasmic reticulum / release of sequestered calcium ion into cytosol / regulation of ryanodine-sensitive calcium-release channel activity / Ion homeostasis / sarcoplasmic reticulum membrane / calcium channel complex / regulation of cytosolic calcium ion concentration / peptidylprolyl isomerase / peptidyl-prolyl cis-trans isomerase activity / response to hydrogen peroxide / Stimuli-sensing channels / Z disc / positive regulation of cytosolic calcium ion concentration / protein refolding / transmembrane transporter binding / signaling receptor binding / membrane / cytosol / cytoplasm
Similarity search - Function
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
Similarity search - Domain/homology
Peptidyl-prolyl cis-trans isomerase FKBP1B
Similarity search - Component
Biological speciesHomo sapiens (human)
Oryctolagus cuniculus (rabbit)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 11 Å
AuthorsDhindwal, S. / Lobo, J.J. / Samso, M.
Funding support United States, 2items
OrganizationGrant numberCountry
American Heart Association14GRNT19660003 United States
Muscular Dystrophy AssociationMDA352845 United States
CitationJournal: Sci Signal / Year: 2017
Title: A cryo-EM-based model of phosphorylation- and FKBP12.6-mediated allosterism of the cardiac ryanodine receptor.
Authors: Sonali Dhindwal / Joshua Lobo / Vanessa Cabra / Demetrio J Santiago / Ashok R Nayak / Kelly Dryden / Montserrat Samsó /
Abstract: Type 2 ryanodine receptors (RyR2s) are calcium channels that play a vital role in triggering cardiac muscle contraction by releasing calcium from the sarcoplasmic reticulum into the cytoplasm. ...Type 2 ryanodine receptors (RyR2s) are calcium channels that play a vital role in triggering cardiac muscle contraction by releasing calcium from the sarcoplasmic reticulum into the cytoplasm. Several cardiomyopathies are associated with the abnormal functioning of RyR2. We determined the three-dimensional structure of rabbit RyR2 in complex with the regulatory protein FKBP12.6 in the closed state at 11.8 Å resolution using cryo-electron microscopy and built an atomic model of RyR2. The heterogeneity in the data set revealed two RyR2 conformations that we proposed to be related to the extent of phosphorylation of the P2 domain. Because the more flexible conformation may correspond to RyR2 with a phosphorylated P2 domain, we suggest that phosphorylation may set RyR2 in a conformation that needs less energy to transition to the open state. Comparison of RyR2 from cardiac muscle and RyR1 from skeletal muscle showed substantial structural differences between the two, especially in the helical domain 2 (HD2) structure forming the Clamp domain, which participates in quaternary interactions with the dihydropyridine receptor and neighboring RyRs in RyR1 but not in RyR2. Rigidity of the HD2 domain of RyR2 was enhanced by binding of FKBP12.6, a ligand that stabilizes RyR2 in the closed state. These results help to decipher the molecular basis of the different mechanisms of activation and oligomerization of the RyR isoforms and could be extended to RyR complexes in other tissues.
History
DepositionJul 29, 2016Deposition site: RCSB / Processing site: RCSB
Revision 1.0May 24, 2017Provider: repository / Type: Initial release
Revision 1.1Jul 18, 2018Group: Data collection / Experimental preparation / Category: em_sample_support / em_software / Item: _em_sample_support.grid_type / _em_software.name
Revision 1.2Mar 6, 2024Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / refine
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _refine.ls_d_res_high / _refine.ls_d_res_low

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Structure visualization

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  • Deposited structure unit
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  • Imaged by UCSF Chimera
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Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Ryanodine Receptor
B: Peptidyl-prolyl cis-trans isomerase FKBP1B
C: Ryanodine Receptor
D: Peptidyl-prolyl cis-trans isomerase FKBP1B
E: Ryanodine Receptor
F: Peptidyl-prolyl cis-trans isomerase FKBP1B
G: Ryanodine Receptor
H: Peptidyl-prolyl cis-trans isomerase FKBP1B


Theoretical massNumber of molelcules
Total (without water)2,019,0688
Polymers2,019,0688
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area34670 Å2
ΔGint-170 kcal/mol
Surface area732100 Å2

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Components

#1: Protein
Ryanodine Receptor /


Mass: 487120.906 Da / Num. of mol.: 4 / Source method: isolated from a natural source / Source: (natural) Oryctolagus cuniculus (rabbit)
#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: 17645.984 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 BL21 (bacteria) / References: UniProt: P68106, peptidylprolyl isomerase

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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

ComponentName: Ryanodine Receptor - FKBP12.6 / Type: COMPLEX / Entity ID: all / Source: MULTIPLE SOURCES
Molecular weightValue: 2.26 MDa / Experimental value: YES
Source (natural)Organism: Oryctolagus cuniculus (rabbit)
Buffer solutionpH: 7.4 / Details: 1x Protease Inhibitor cocktail
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMMOPS sodiumC7H14NNaO4S1
2200 mMsodium chlorideNaClSodium chloride1
32 mMdithiothreitolC4H10O2S21
40.015 %Tween 20(Polyoxyethylene (20) sorbitan monolaurate)C58H114O261
52 mMEGTAC14H24N2O101
SpecimenConc.: 0.1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 295.15 K
Details: Blot for 2 seconds before plunging into liquid ethane (FEI VITROBOT MARK IV).

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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 FIELDBright-field microscopy / Nominal magnification: 59000 X / Calibrated magnification: 62000 X / Nominal defocus max: 6000 nm / Nominal defocus min: 2000 nm / Cs: 2.7 mm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 1.2 sec. / Electron dose: 20 e/Å2 / Detector mode: OTHER / Film or detector model: FEI FALCON II (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 858
Image scansSampling size: 15 µm / Width: 4096 / Height: 4096 / Movie frames/image: 7 / Used frames/image: 1-7

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Processing

SoftwareName: REFMAC / Version: 5.8.0135 / Classification: refinement
EM software
IDNameVersionCategory
1RELION1.3particle selection
2EPUimage acquisition
4CTFFIND3.5CTF correction
5FREALIGN9.11CTF correction
8Situs2.8model fitting
9UCSF Chimera1.11model fitting
11REFMAC5.8model refinement
12FREALIGN9.11initial Euler assignment
13FREALIGN9.11final Euler assignment
15FREALIGN9.113D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 13600
Details: Used Relion 1.3 to auto-pick the particles. Inspected each micrograph for false positives.
SymmetryPoint symmetry: C4 (4 fold cyclic)
3D reconstructionResolution: 11 Å / Resolution method: FSC 0.5 CUT-OFF / Num. of particles: 13158 / Algorithm: FOURIER SPACE / Details: FREALIGN half maps were used to calculate FSC. / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: RECIPROCAL
RefinementResolution: 11→11 Å / Cor.coef. Fo:Fc: 0.976 / SU B: 505.233 / SU ML: 2.74
Stereochemistry target values: MAXIMUM LIKELIHOOD WITH PHASES
Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
RfactorNum. reflection% reflection
Rwork0.28217 --
obs0.28217 57758 100 %
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK
Displacement parametersBiso mean: 461.311 Å2
Baniso -1Baniso -2Baniso -3
1-1.97 Å20.86 Å21.15 Å2
2--2.17 Å2-0.75 Å2
3----4.15 Å2
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
ELECTRON MICROSCOPYr_bond_refined_d0.0160.019115032
ELECTRON MICROSCOPYr_bond_other_d0.0020.02110156
ELECTRON MICROSCOPYr_angle_refined_deg1.2811.948155516
ELECTRON MICROSCOPYr_angle_other_deg1.0443252220
ELECTRON MICROSCOPYr_dihedral_angle_1_deg6.52514452
ELECTRON MICROSCOPYr_dihedral_angle_2_deg31.54823.9565136
ELECTRON MICROSCOPYr_dihedral_angle_3_deg16.0381518904
ELECTRON MICROSCOPYr_dihedral_angle_4_deg11.31615688
ELECTRON MICROSCOPYr_chiral_restr0.0650.217544
ELECTRON MICROSCOPYr_gen_planes_refined0.0040.02130708
ELECTRON MICROSCOPYr_gen_planes_other0.0010.0226828
ELECTRON MICROSCOPYr_nbd_refined
ELECTRON MICROSCOPYr_nbd_other
ELECTRON MICROSCOPYr_nbtor_refined
ELECTRON MICROSCOPYr_nbtor_other
ELECTRON MICROSCOPYr_xyhbond_nbd_refined
ELECTRON MICROSCOPYr_xyhbond_nbd_other
ELECTRON MICROSCOPYr_metal_ion_refined
ELECTRON MICROSCOPYr_metal_ion_other
ELECTRON MICROSCOPYr_symmetry_vdw_refined
ELECTRON MICROSCOPYr_symmetry_vdw_other
ELECTRON MICROSCOPYr_symmetry_hbond_refined
ELECTRON MICROSCOPYr_symmetry_hbond_other
ELECTRON MICROSCOPYr_symmetry_metal_ion_refined
ELECTRON MICROSCOPYr_symmetry_metal_ion_other
ELECTRON MICROSCOPYr_mcbond_it6.40846.06858576
ELECTRON MICROSCOPYr_mcbond_other6.40846.06858575
ELECTRON MICROSCOPYr_mcangle_it11.28269.1972772
ELECTRON MICROSCOPYr_mcangle_other11.28269.1972773
ELECTRON MICROSCOPYr_scbond_it3.29447.13456456
ELECTRON MICROSCOPYr_scbond_other3.29447.13456454
ELECTRON MICROSCOPYr_scangle_it
ELECTRON MICROSCOPYr_scangle_other6.88570.49182744
ELECTRON MICROSCOPYr_long_range_B_refined26.965309865
ELECTRON MICROSCOPYr_long_range_B_other26.965309866
ELECTRON MICROSCOPYr_rigid_bond_restr
ELECTRON MICROSCOPYr_sphericity_free
ELECTRON MICROSCOPYr_sphericity_bonded
LS refinement shellResolution: 10.5→10.772 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rwork0.391 4283 -
Rfree-0 -
obs--100 %

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