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- EMDB-45497: Rabbit RyR1 disease mutant Y523S in complex with FKBP12.6, nanodi... -

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Entry
Database: EMDB / ID: EMD-45497
TitleRabbit RyR1 disease mutant Y523S in complex with FKBP12.6, nanodisc and inhibitor dantrolene in the presence of activating calcium
Map data
Sample
  • Complex: Rabbit RyR1 disease mutant Y523S in complex with FKBP12.6, nanodisc and inhibitor dantrolene in the presence of activating calcium
    • Protein or peptide: RyR1 Y523S
    • Protein or peptide: FKBP12.6
KeywordsRyanodine receptor / Calcium channel / Mutation / Y523S / RyR1 / dantrolene / MEMBRANE PROTEIN / MEMBRANE PROTEIN-INHIBITOR complex
Biological speciesOryctolagus cuniculus (rabbit) / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.23 Å
AuthorsIyer KA / Samso M
Funding support United States, 5 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIH/NIAMS)R01 AR068431 United States
American Heart Association19POST34430178 United States
Other privateMDA 352845
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)U24 GM116789 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)U24 GM116790 United States
CitationJournal: Structure / Year: 2025
Title: Dantrolene inhibition of ryanodine receptor 1 carrying the severe malignant hyperthermia mutation Y522S visualized by cryo-EM.
Authors: Kavita A Iyer / Takuya Kobayashi / Takashi Murayama / Montserrat Samsó /
Abstract: Mutations in the skeletal isoform of the ryanodine receptor 1 (RyR1) pose grave risks during anesthesia or treatment with succinylcholine muscle relaxants. These can trigger a potentially lethal ...Mutations in the skeletal isoform of the ryanodine receptor 1 (RyR1) pose grave risks during anesthesia or treatment with succinylcholine muscle relaxants. These can trigger a potentially lethal malignant hyperthermia (MH) episode via intracellular calcium increase mainly from RyR1 channel leakage. Dantrolene is the only known treatment option to prevent death. The main target of dantrolene is RyR1; however, little is known about the mechanism of inhibition. Cryoelectron microscopy (cryo-EM) structures of dantrolene bound to the severe MH Y522S RyR1 mutant in the closed and open states at 2.5-3.3 Å resolution revealed that the drug binds to the channel's cytoplasmic assembly, far from the ion gate, interacting with residues W882, W996, and R1000 in the P1 domain. The finding was validated by Ca imaging and [H]ryanodine binding in wild-type (WT) and alanine mutants. Dantrolene reduced channel opening probability by restricting the central activation module, "cooling down" the primed conformation caused by the mutation.
History
DepositionJun 26, 2024-
Header (metadata) releaseDec 11, 2024-
Map releaseDec 11, 2024-
UpdateFeb 19, 2025-
Current statusFeb 19, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_45497.png

Downloads & links

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Map

FileDownload / File: emd_45497.map.gz / Format: CCP4 / Size: 381.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.08 Å/pix.
x 464 pix.
= 501.12 Å		1.08 Å/pix.
x 464 pix.
= 501.12 Å		1.08 Å/pix.
x 464 pix.
= 501.12 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.08 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.335
Minimum - Maximum-1.9749615 - 3.9202394
Average (Standard dev.)0.004272291 (±0.09509099)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions464464464
Spacing464464464
CellA=B=C: 501.12003 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_45497_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_45497_half_map_2.map
Projections & Slices
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Histogram (log scale)

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Sample components

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Entire : Rabbit RyR1 disease mutant Y523S in complex with FKBP12.6, nanodi...

EntireName: Rabbit RyR1 disease mutant Y523S in complex with FKBP12.6, nanodisc and inhibitor dantrolene in the presence of activating calcium
Components
  • Complex: Rabbit RyR1 disease mutant Y523S in complex with FKBP12.6, nanodisc and inhibitor dantrolene in the presence of activating calcium
    • Protein or peptide: RyR1 Y523S
    • Protein or peptide: FKBP12.6

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Supramolecule #1: Rabbit RyR1 disease mutant Y523S in complex with FKBP12.6, nanodi...

SupramoleculeName: Rabbit RyR1 disease mutant Y523S in complex with FKBP12.6, nanodisc and inhibitor dantrolene in the presence of activating calcium
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Oryctolagus cuniculus (rabbit)
Molecular weightTheoretical: 2.26 MDa

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Macromolecule #1: RyR1 Y523S

MacromoleculeName: RyR1 Y523S / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Oryctolagus cuniculus (rabbit)
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MGDGGEGEDE VQFLRTDDEV VLQCSATVLK EQLKLCLAAE GFGNRLCFLE PTSNAQNVPP DLAICCFTL EQSLSVRALQ EMLANTVEAG VESSQGGGHR TLLYGHAILL RHAHSRMYLS C LTTSRSMT DKLAFDVGLQ EDATGEACWW TMHPASKQRS EGEKVRVGDD ...String:
MGDGGEGEDE VQFLRTDDEV VLQCSATVLK EQLKLCLAAE GFGNRLCFLE PTSNAQNVPP DLAICCFTL EQSLSVRALQ EMLANTVEAG VESSQGGGHR TLLYGHAILL RHAHSRMYLS C LTTSRSMT DKLAFDVGLQ EDATGEACWW TMHPASKQRS EGEKVRVGDD LILVSVSSER YL HLSTASG ELQVDASFMQ TLWNMNPICS CCEEGYVTGG HVLRLFHGHM DECLTISAAD SDD QRRLVY YEGGAVCTHA RSLWRLEPLR ISWSGSHLRW GQPLRIRHVT TGRYLALTED QGLV VVDAC KAHTKATSFC FRVSKEKLDT APKRDVEGMG PPEIKYGESL CFVQHVASGL WLTYA APDP KALRLGVLKK KAILHQEGHM DDALFLTRCQ QEESQAARMI HSTAGLYNQF IKGLDS FSG KPRGSGPPAG PALPIEAVIL SLQDLIGYFE PPSEELQHEE KQSKLRSLRN RQSLFQE EG MLSLVLNCID RLNVYTTAAH FAEYAGEEAA ESWKEIVNLL SELLASLIRG NRANCALF S TNLDWVVSKL DRLEASSGIL EVLYCVLIES PEVLNIIQEN HIKSIISLLD KHGRNHKVL DVLCSLCVCN GVAVRSNQDL ITENLLPGRE LLLQTNLINY VTSIRPNIFV GRAEGSTQYG KWYFEVMVD EVVPFLTAQA THLRVGWALT EGYSPYPGGG EGWGGNGVGD DLYSYGFDGL H LWTGHVAR PVTSPGQHLL APEDVVSCCL DLSVPSISFR INGCPVQGVF EAFNLDGLFF PV VSFSAGV KVRFLLGGRH GEFKFLPPPG YAPCHEAVLP RERLRLEPIK EYRREGPRGP HLV GPSRCL SHTDFVPCPV DTVQIVLPPH LERIREKLAE NIHELWALTR IEQGWTYGPV RDDN KRLHP CLVNFHSLPE PERNYNLQMS GETLKTLLAL GCHVGMADEK AEDNLKKTKL PKTYM MSNG YKPAPLDLSH VRLTPAQTTL VDRLAENGHN VWARDRVAQG WSYSAVQDIP ARRNPR LVP YRLLDEATKR SNRDSLCQAV RTLLGYGYNI EPPDQEPSQV ENQSRWDRVR IFRAEKS YT VQSGRWYFEF EAVTTGEMRV GWARPELRPD VELGADELAY VFNGHRGQRW HLGSEPFG R PWQSGDVVGC MIDLTENTII FTLNGEVLMS DSGSETAFRE IEIGDGFLPV CSLGPGQVG HLNLGQDVSS LRFFAICGLQ EGFEPFAINM QRPVTTWFSK SLPQFEPVPP EHPHYEVARM DGTVDTPPC LRLAHRTWGS QNSLVEMLFL RLSLPVQFHQ HFRCTAGATP LAPPGLQPPA E DEARAAEP DPDYENLRRS AGGWGEAEGG KEGTAKEGTP GGTPQPGVEA QPVRAENEKD AT TEKNKKR GFLFKAKKAA MMTQPPATPA LPRLPHDVVP ADNRDDPEII LNTTTYYYSV RVF AGQEPS CVWVGWVTPD YHQHDMNFDL SKVRAVTVTM GDEQGNVHSS LKCSNCYMVW GGDF VSPGQ QGRISHTDLV IGCLVDLATG LMTFTANGKE SNTFFQVEPN TKLFPAVFVL PTHQN VIQF ELGKQKNIMP LSAAMFLSER KNPAPQCPPR LEVQMLMPVS WSRMPNHFLQ VETRRA GER LGWAVQCQDP LTMMALHIPE ENRCMDILEL SERLDLQRFH SHTLRLYRAV CALGNNR VA HALCSHVDQA QLLHALEDAH LPGPLRAGYY DLLISIHLES ACRSRRSMLS EYIVPLTP E TRAITLFPPG RKGGNARRHG LPGVGVTTSL RPPHHFSPPC FVAALPAAGV AEAPARLSP AIPLEALRDK ALRMLGEAVR DGGQHARDPV GGSVEFQFVP VLKLVSTLLV MGIFGDEDVK QILKMIEPE VFTEEEEEEE EEEEEEEEEE EDEEEKEEDE EEEEKEDAEK EEEEAPEGEK E DLEEGLLQ MKLPESVKLQ MCNLLEYFCD QELQHRVESL AAFAERYVDK LQANQRSRYA LL MRAFTMS AAETARRTRE FRSPPQEQIN MLLHFKDEAD EEDCPLPEDI RQDLQDFHQD LLA HCGIQL EGEEEEPEEE TSLSSRLRSL LETVRLVKKK EEKPEEELPA EEKKPQSLQE LVSH MVVRW AQEDYVQSPE LVRAMFSLLH RQYDGLGELL RALPRAYTIS PSSVEDTMSL LECLG QIRS LLIVQMGPQE ENLMIQSIGN IMNNKVFYQH PNLMRALGMH ETVMEVMVNV LGGGET KEI RFPKMVTSCC RFLCYFCRIS RQNQRSMFDH LSYLLENSGI GLGMQGSTPL DVAAASV ID NNELALALQE QDLEKVVSYL AGCGLQSCPM LLAKGYPDIG WNPCGGERYL DFLRFAVF V NGESVEENAN VVVRLLIRKP ECFGPALRGE GGSGLLAAIE EAIRISEDPA RDGPGVRRD RRREHFGEEP PEENRVHLGH AIMSFYAALI DLLGRCAPEM HLIQAGKGEA LRIRAILRSL VPLDDLVGI ISLPLQIPTL GKDGALVQPK MSASFVPDHK ASMVLFLDRV YGIENQDFLL H VLDVGFLP DMRAAASLDT ATFSTTEMAL ALNRYLCLAV LPLITKCAPL FAGTEHRAIM VD SMLHTVY RLSRGRSLTK AQRDVIEDCL MALCRYIRPS MLQHLLRRLV FDVPILNEFA KMP LKLLTN HYERCWKYYC LPTGWANFGV TSEEELHLTR KLFWGIFDSL AHKKYDQELY RMAM PCLCA IAGALPPDYV DASYSSKAEK KATVDAEGNF DPRPVETLNV IIPEKLDSFI NKFAE YTHE KWAFDKIQNN WSYGENVDEE LKTHPMLRPY KTFSEKDKEI YRWPIKESLK AMIAWE WTI EKAREGEEER TEKKKTRKIS QTAQTYDPRE GYNPQPPDLS GVTLSRELQA MAEQLAE NY HNTWGRKKKQ ELEAKGGGTH PLLVPYDTLT AKEKARDREK AQELLKFLQM NGYAVTRG L KDMELDTSSI EKRFAFGFLQ QLLRWMDISQ EFIAHLEAVV SSGRVEKSPH EQEIKFFAK ILLPLINQYF TNHCLYFLST PAKVLGSGGH ASNKEKEMIT SLFCKLAALV RHRVSLFGTD APAVVNCLH ILARSLDART VMKSGPEIVK AGLRSFFESA SEDIEKMVEN LRLGKVSQAR T QVKGVGQN LTYTTVALLP VLTTLFQHIA QHQFGDDVIL DDVQVSCYRT LCSIYSLGTT KN TYVEKLR PALGECLARL AAAMPVAFLE PQLNEYNACS VYTTKSPRER AILGLPNSVE EMC PDIPVL DRLMADIGGL AESGARYTEM PHVIEITLPM LCSYLPRWWE RGPEAPPPAL PAGA PPPCT AVTSDHLNSL LGNILRIIVN NLGIDEATWM KRLAVFAQPI VSRARPELLH SHFIP TIGR LRKRAGKVVA EEEQLRLEAK AEAEEGELLV RDEFSVLCRD LYALYPLLIR YVDNNR AHW LTEPNANAEE LFRMVGEIFI YWSKSHNFKR EEQNFVVQNE INNMSFLTAD SKSKMAK AG DAQSGGSDQE RTKKKRRGDR YSVQTSLIVA TLKKMLPIGL NMCAPTDQDL IMLAKTRY A LKDTDEEVRE FLQNNLHLQG KVEGSPSLRW QMALYRGLPG REEDADDPEK IVRRVQEVS AVLYHLEQTE HPYKSKKAVW HKLLSKQRRR AVVACFRMTP LYNLPTHRAC NMFLESYKAA WILTEDHSF EDRMIDDLSK AGEQEEEEEE VEEKKPDPLH QLVLHFSRTA LTEKSKLDED Y LYMAYADI MAKSCHLEEG GENGEAEEEE VEVSFEEKEM EKQRLLYQQS RLHTRGAAEM VL QMISACK GETGAMVSST LKLGISILNG GNAEVQQKML DYLKDKKEVG FFQSIQALMQ TCS VLDLNA FERQNKAEGL GMVNEDGTVI NRQNGEKVMA DDEFTQDLFR FLQLLCEGHN NDFQ NYLRT QTGNTTTINI IICTVDYLLR LQESISDFYW YYSGKDVIEE QGKRNFSKAM SVAKQ VFNS LTEYIQGPCT GNQQSLAHSR LWDAVVGFLH VFAHMMMKLA QDSSQIELLK ELLDLQ KDM VVMLLSLLEG NVVNGMIARQ MVDMLVESSS NVEMILKFFD MFLKLKDIVG SEAFQDY VT DPRGLISKKD FQKAMDSQKQ FTGPEIQFLL SCSEADENEM INFEEFANRF QEPARDIG F NVAVLLTNLS EHVPHDPRLR NFLELAESIL EYFRPYLGRI EIMGASRRIE RIYFEISET NRAQWEMPQV KESKRQFIFD VVNEGGEAEK MELFVSFCED TIFEMQIAAQ ISEPEGEPEA DEDEGMGEA AAEGAEEGAA GAEGAAGTVA AGATARLAAA AARALRGLSY RSLRRRVRRL R RLTAREAA TALAALLWAV VARAGAAGAG AAAGALRLLW GSLFGGGLVE GAKKVTVTEL LA GMPDPTS DEVHGEQPAG PGGDADGAGE GEGEGDAAEG DGDEEVAGHE AGPGGAEGVV AVA DGGPFR PEGAGGLGDM GDTTPAEPPT PEGSPILKRK LGVDGEEEEL VPEPEPEPEP EPEK ADEEN GEKEEVPEAP PEPPKKAPPS PPAKKEEAGG AGMEFWGELE VQRVKFLNYL SRNFY TLRF LALFLAFAIN FILLFYKVSD SPPGEDDMEG SAAGDLAGAG SGGGSGWGSG AGEEAE GDE DENMVYYFLE ESTGYMEPAL WCLSLLHTLV AFLCIIGYNC LKVPLVIFKR EKELARK LE FDGLYITEQP GDDDVKGQWD RLVLNTPSFP SNYWDKFVKR KVLDKHGDIF GRERIAEL L GMDLASLEIT AHNERKPDPP PGLLTWLMSI DVKYQIWKFG VIFTDNSFLY LGWYMVMSL LGHYNNFFFA AHLLDIAMGV KTLRTILSSV THNGKQLVMT VGLLAVVVYL YTVVAFNFFR KFYNKSEDE DEPDMKCDDM MTCYLFHMYV GVRAGGGIGD EIEDPAGDEY ELYRVVFDIT F FFFVIVIL LAIIQGLIID AFGELRDQQE QVKEDMETKC FICGIGSDYF DTTPHGFETH TL EEHNLAN YMFFLMYLIN KDETEHTGQE SYVWKMYQER CWDFFPAGDC FRKQYEDQLS

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Macromolecule #2: FKBP12.6

MacromoleculeName: FKBP12.6 / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
SequenceString:
GVEIETISPG DGRTFPKKG Q TCVVHYTG ML QNGKKFD SSR DRNKPF KFRI GKQEV IKGFE EGAA QMSLGQ RAK LTCTPDV AY GATGHPGV I PPNATLIFD VELLNLE

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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

Concentration3 mg/mL
BufferpH: 7.4
Component:
ConcentrationFormulaName
20.0 mMMOPS3-(N-Morpholino)propanesulfonic acid sodium salt
0.7 MNaClSodium Chloride
2.0 mMDTTDithiothreitol
2.0 mMATPAdenosine 5-triphosphate
0.825 mMCaCl2Calcium Chloride
0.5 mMEGTAEthylene glycol-bis(beta-aminoethyl ether)-N,N,N,N-tetraacetic acid
0.05 mMDantroleneDantrolene
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Slit width: 10 eV
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 2 / Number real images: 12212 / Average exposure time: 2.69 sec. / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.6 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 81000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1968209
Startup modelType of model: NONE
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C4 (4 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.23 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.3.1) / Number images used: 189847
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3.1)
Final angle assignmentType: PROJECTION MATCHING / Software - Name: cryoSPARC (ver. 3.3.1)
Final 3D classificationNumber classes: 4 / Software - Name: cryoSPARC (ver. 3.3.1)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

7t65


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: RIGID BODY FIT

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