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- EMDB-14120: Human RZZ kinetochore corona complex. -

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Basic information

Entry
Database: EMDB / ID: EMD-14120
TitleHuman RZZ kinetochore corona complex.
Map dataComposite map from focussed refinement maps
Sample
  • Complex: RZZ complex
    • Protein or peptide: Protein zwilch homolog
    • Protein or peptide: Kinetochore-associated protein 1
    • Protein or peptide: Centromere/kinetochore protein zw10 homolog
Function / homology
Function and homology information


protein localization to kinetochore involved in kinetochore assembly / RZZ complex / Dsl1/NZR complex / kinetochore microtubule / centromeric DNA binding / regulation of attachment of spindle microtubules to kinetochore / regulation of exit from mitosis / protein localization to kinetochore / COPI-dependent Golgi-to-ER retrograde traffic / mitotic metaphase chromosome alignment ...protein localization to kinetochore involved in kinetochore assembly / RZZ complex / Dsl1/NZR complex / kinetochore microtubule / centromeric DNA binding / regulation of attachment of spindle microtubules to kinetochore / regulation of exit from mitosis / protein localization to kinetochore / COPI-dependent Golgi-to-ER retrograde traffic / mitotic metaphase chromosome alignment / mitotic spindle assembly checkpoint signaling / Golgi organization / establishment of mitotic spindle orientation / mitotic sister chromatid segregation / Amplification of signal from unattached kinetochores via a MAD2 inhibitory signal / endoplasmic reticulum to Golgi vesicle-mediated transport / Mitotic Prometaphase / EML4 and NUDC in mitotic spindle formation / Resolution of Sister Chromatid Cohesion / lipid droplet / bioluminescence / meiotic cell cycle / generation of precursor metabolites and energy / RHO GTPases Activate Formins / kinetochore / small GTPase binding / spindle pole / Separation of Sister Chromatids / protein transport / actin cytoskeleton / protein-containing complex assembly / cell division / endoplasmic reticulum membrane / endoplasmic reticulum / membrane / nucleus / plasma membrane / cytoplasm / cytosol
Similarity search - Function
RZZ complex, subunit Zw10 / RZZ complex, subunit KNTC1/ROD, C-terminal / Centromere/kinetochore Zw10 N-terminal / Rough deal protein C-terminal region / RZZ complex, subunit Zwilch / RZZ complex, subunit zwilch / Green fluorescent protein, GFP / Green fluorescent protein-related / Green fluorescent protein / Green fluorescent protein / WD40-repeat-containing domain superfamily
Similarity search - Domain/homology
Uncharacterized protein / Centromere/kinetochore protein zw10 homolog / Kinetochore-associated protein 1 / Protein zwilch homolog
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.9 Å
AuthorsRaisch T / Ciossani G / d'Amico E / Cmetowski V / Carmignani S / Maffini S / Merino F / Wohlgemuth S / Vetter IR / Raunser S / Musacchio A
Funding support Germany, 1 items
OrganizationGrant numberCountry
Max Planck Society Germany
CitationJournal: EMBO J / Year: 2022
Title: Structure of the RZZ complex and molecular basis of Spindly-driven corona assembly at human kinetochores.
Authors: Tobias Raisch / Giuseppe Ciossani / Ennio d'Amico / Verena Cmentowski / Sara Carmignani / Stefano Maffini / Felipe Merino / Sabine Wohlgemuth / Ingrid R Vetter / Stefan Raunser / Andrea Musacchio /
Abstract: In metazoans, a ≈1 megadalton (MDa) multiprotein complex comprising the dynein-dynactin adaptor Spindly and the ROD-Zwilch-ZW10 (RZZ) complex is the building block of a fibrous biopolymer, the ...In metazoans, a ≈1 megadalton (MDa) multiprotein complex comprising the dynein-dynactin adaptor Spindly and the ROD-Zwilch-ZW10 (RZZ) complex is the building block of a fibrous biopolymer, the kinetochore fibrous corona. The corona assembles on mitotic kinetochores to promote microtubule capture and spindle assembly checkpoint (SAC) signaling. We report here a high-resolution cryo-EM structure that captures the essential features of the RZZ complex, including a farnesyl-binding site required for Spindly binding. Using a highly predictive in vitro assay, we demonstrate that the SAC kinase MPS1 is necessary and sufficient for corona assembly at supercritical concentrations of the RZZ-Spindly (RZZS) complex, and describe the molecular mechanism of phosphorylation-dependent filament nucleation. We identify several structural requirements for RZZS polymerization in rings and sheets. Finally, we identify determinants of kinetochore localization and corona assembly of Spindly. Our results describe a framework for the long-sought-for molecular basis of corona assembly on metazoan kinetochores.
History
DepositionJan 4, 2022-
Header (metadata) releaseMar 16, 2022-
Map releaseMar 16, 2022-
UpdateMay 11, 2022-
Current statusMay 11, 2022Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.4
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.4
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7qpg
  • Surface level: 0.4
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-7qpg
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_14120.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationComposite map from focussed refinement maps
Voxel sizeX=Y=Z: 1.8 Å
Density
Contour LevelBy AUTHOR: 0.4 / Movie #1: 0.4
Minimum - Maximum0.0 - 3.6767004
Average (Standard dev.)0.003982301 (±0.050687198)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 540.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.81.81.8
M x/y/z300300300
origin x/y/z0.0000.0000.000
length x/y/z540.000540.000540.000
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ450450450
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS300300300
D min/max/mean0.0003.6770.004

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Supplemental data

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Mask #1

Fileemd_14120_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Mask #2

Fileemd_14120_msk_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: Map from global Non-uniform refinement

Fileemd_14120_additional_1.map
AnnotationMap from global Non-uniform refinement
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: Locally filtered map

Fileemd_14120_additional_2.map
AnnotationLocally filtered map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire : RZZ complex

EntireName: RZZ complex
Components
  • Complex: RZZ complex
    • Protein or peptide: Protein zwilch homolog
    • Protein or peptide: Kinetochore-associated protein 1
    • Protein or peptide: Centromere/kinetochore protein zw10 homolog

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Supramolecule #1: RZZ complex

SupramoleculeName: RZZ complex / type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)

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Macromolecule #1: Protein zwilch homolog

MacromoleculeName: Protein zwilch homolog / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 67.293805 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString: MWERLNCAAE DFYSRLLQKF NEEKKGIRKD PFLYEADVQV QLISKGQPNP LKNILNENDI VFIVEKVPLE KEETSHIEEL QSEETAISD FSTGENVGPL ALPVGKARQL IGLYTMAHNP NMTHLKINLP VTALPPLWVR CDSSDPEGTC WLGAELITTN N SITGIVLY ...String:
MWERLNCAAE DFYSRLLQKF NEEKKGIRKD PFLYEADVQV QLISKGQPNP LKNILNENDI VFIVEKVPLE KEETSHIEEL QSEETAISD FSTGENVGPL ALPVGKARQL IGLYTMAHNP NMTHLKINLP VTALPPLWVR CDSSDPEGTC WLGAELITTN N SITGIVLY VVSCKADKNY SVNLENLKNL HKKRHHLSTV TSKGFAQYEL FKSSALDDTI TASQTAIALD ISWSPVDEIL QI PPLSSTA TLNIKVESGE PRGPLNHLYR ELKFLLVLAD GLRTGVTEWL EPLEAKSAVE LVQEFLNDLN KLDGFGDSTK KDT EVETLK HDTAAVDRSV KRLFKVRSDL DFAEQLWCKM SSSVISYQDL VKCFTLIIQS LQRGDIQPWL HSGSNSLLSK LIHQ SYHGT MDTVSLSGTI PVQMLLEIGL DKLKKDYISF FIGQELASLN HLEYFIAPSV DIQEQVYRVQ KLHHILEILV SCMPF IKSQ HELLFSLTQI CIKYYKQNPL DEQHIFQLPV RPTAVKNLYQ SEKPQKWRVE IYSGQKKIKT VWQLSDSSPI DHLNFH KPD FSELTLNGSL EERIFFTNMV TCSQVHFK

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Macromolecule #2: Kinetochore-associated protein 1

MacromoleculeName: Kinetochore-associated protein 1 / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 279.805562 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString: MAHHHHHHSS GVSKGEEDNM AIIKEFMRFK VHMEGSVNGH EFEIEGEGEG RPYEGTQTAK LKVTKGGPLP FAWDILSPQF MYGSKAYVK HPADIPDYLK LSFPEGFKWE RVMNFEDGGV VTVTQDSSLQ DGEFIYKVKL RGTNFPSDGP VMQKKTMGWE A SSERMYPE ...String:
MAHHHHHHSS GVSKGEEDNM AIIKEFMRFK VHMEGSVNGH EFEIEGEGEG RPYEGTQTAK LKVTKGGPLP FAWDILSPQF MYGSKAYVK HPADIPDYLK LSFPEGFKWE RVMNFEDGGV VTVTQDSSLQ DGEFIYKVKL RGTNFPSDGP VMQKKTMGWE A SSERMYPE DGALKGEIKQ RLKLKDGGHY DAEVKTTYKA KKPVQLPGAY NVNIKLDITS HNEDYTIVEQ YERAEGRHST GG MDELYKL EVLFQGPGSW NDIELLTNDD TGSGYLSVGS RKEHGTALYQ VDLLVKISSE KASLNPKIQA CSLSDGFIIV ADQ SVILLD SICRSLQLHL VFDTEVDVVG LCQEGKFLLV GERSGNLHLI HVTSKQTLLT NAFVQKANDE NRRTYQNLVI EKDG SNEGT YYMLLLTYSG FFCITNLQLL KIQQAIENVD FSTAKKLQGQ IKSSFISTEN YHTLGCLSLV AGDLASEVPV IIGGT GNCA FSKWEPDSSK KGMTVKNLID AEIIKGAKKF QLIDNLLFVL DTDNVLSLWD IYTLTPVWNW PSLHVEEFLL TTEADS PSS VTWQGITNLK LIALTASANK KMKNLMVYSL PTMEILYSLE VSSVSSLVQT GISTDTIYLL EGVCKNDPKL SEDSVSV LV LRCLTEALPE NRLSRLLHKH RFAEAESFAI QFGLDVELVY KVKSNHILEK LALSSVDASE QTEWQQLVDD AKENLHKI Q DDEFVVNYCL KAQWITYETT QEMLNYAKTR LLKKEDKTAL IYSDGLKEVL RAHAKLTTFY GAFGPEKFSG SSWIEFLNN EDDLKDIFLQ LKEGNLVCAQ YLWLRHRANF ESRFDVKMLE SLLNSMSASV SLQKLCPWFK NDVIPFVRRT VPEGQIILAK WLEQAARNL ELTDKANWPE NGLQLAEIFF TAEKTDELGL ASSWHWISLK DYQNTEEVCQ LRTLVNNLRE LITLHRKYNC K LALSDFEK ENTTTIVFRM FDKVLAPELI PSILEKFIRV YMREHDLQEE ELLLLYIEDL LNRCSSKSTS LFETAWEAKA MA VIACLSD TDLIFDAVLK IMYAAVVPWS AAVEQLVKQH LEMDHPKVKL LQESYKLMEM KKLLRGYGIR EVNLLNKEIM RVV RYILKQ DVPSSLEDAL KVAQAFMLSD DEIYSLRIID LIDREQGEDC LLLLKSLPPA EAEKTAERVI IWARLALQEE PDHS KEGKA WRMSVAKTSV DILKILCDIQ KDNLQKKDEC EEMLKLFKEV ASLQENFEVF LSFEDYSNSS LVADLREQHI KAHEV AQAK HKPGSTPEPI AAEVRSPSME SKLHRQALAL QMSKQELEAE LTLRALKDGN IKTALKKCSD LFKYHCNADT GKLLFL TCQ KLCQMLADNV PVTVPVGLNL PSMIHDLASQ AATICSPDFL LDALELCKHT LMAVELSRQC QMDDCGILMK ASFGTHK DP YEEWSYSDFF SEDGIVLESQ MVLPVIYELI SSLVPLAESK RYPLESTSLP YCSLNEGDGL VLPVINSISA LLQNLQES S QWELALRFVV GSFGTCLQHS VSNFMNATLS EKLFGETTLV KSRHVVMELK EKAVIFIREN ATTLLHKVFN CRLVDLDLA LGYCTLLPQK DVFENLWKLI DKAWQNYDKI LAISLVGSEL ASLYQEIEMG LKFRELSTDA QWGIRLGKLG ISFQPVFRQH FLTKKDLIK ALVENIDMDT SLILEYCSTF QLDCDAVLQL FIETLLHNTN AGQGQGDASM DSAKRRHPKL LAKALEMVPL L TSTKDLVI SLSGILHKLD PYDYEMIEVV LKVIERADEK ITNININQAL SILKHLKSYR RISPPVDLEY QYMLEHVITL PS AAQTRLP FHLIFFGTAQ NFWKILSTEL SEESFPTLLL ISKLMKFSLD TLYVSTAKHV FEKKLKPKLL KLTQAKSSTL INK EITKIT QTIESCLLSI VNPEWAVAIA ISLAQDIPEG SFKISALKFC LYLAERWLQN IPSQDEKREK AEALLKKLHI QYRR SGTEA VLIAHKLNTE EYLRVIGKPA HLIVSLYEHP SINQRIQNSS GTDYPDIHAA AKEIAEVNEI NLEKVWDMLL EKWLC PSTK PGEKPSELFE LQEDEALRRV QYLLLSRPID YSSRMLFVFA TSTTTTLGMH QLTFAHRTRA LQCLFYLADK ETIESL FKK PIEEVKSYLR CITFLASFET LNIPITYELF CSSPKEGMIK GLWKNHSHES MAVRLVTELC LEYKIYDLQL WNGLLQK LL GFNMIPYLRK VLKAISSIHS LWQVPYFSKA WQRVIQIPLL SASCPLSPDQ LSDCSESLIA VLECPVSGDL DLIGVARQ Y IQLELPAFAL ACLMLMPHSE KRHQQIKNFL GSCDPQVILK QLEEHMNTGQ LAGFSHQIRS LILNNIINKK EFGILAKTK YFQMLKMHAM NTNNITELVN YLANDLSLDE ASVLITEYSK HCGKPVPPDT APCEILKMFL SGLS

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Macromolecule #3: Centromere/kinetochore protein zw10 homolog

MacromoleculeName: Centromere/kinetochore protein zw10 homolog / type: protein_or_peptide / ID: 3 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 88.940336 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString: MASFVTEVLA HSGRLEKEDL GTRISRLTRR VEEIKGEVCN MISKKYSEFL PSMQSAQGLI TQVDKLSEDI DLLKSRIESE VRRDLHVST GEFTDLKQQL ERDSVVLSLL KQLQEFSTAI EEYNCALTEK KYVTGAQRLE EAQKCLKLLK SRKCFDLKIL K SLSMELTI ...String:
MASFVTEVLA HSGRLEKEDL GTRISRLTRR VEEIKGEVCN MISKKYSEFL PSMQSAQGLI TQVDKLSEDI DLLKSRIESE VRRDLHVST GEFTDLKQQL ERDSVVLSLL KQLQEFSTAI EEYNCALTEK KYVTGAQRLE EAQKCLKLLK SRKCFDLKIL K SLSMELTI QKQNILYHLG EEWQKLIVWK FPPSKDTSSL ESYLQTELHL YTEQSHKEEK TPMPPISSVL LAFSVLGELH SK LKSFGQM LLKYILRPLA SCPSLHAVIE SQPNIVIIRF ESIMTNLEYP SPSEVFTKIR LVLEVLQKQL LDLPLDTDLE NEK TSTVPL AEMLGDMIWE DLSECLIKNC LVYSIPTNSS KLQQYEEIIQ STEEFENALK EMRFLKGDTT DLLKYARNIN SHFA NKKCQ DVIVAARNLM TSEIHNTVKI IPDSKINVPE LPTPDEDNKL EVQKVSNTQY HEVMNLEPEN TLDQHSFSLP TCRIS ESVK KLMELAYQTL LEATTSSDQC AVQLFYSVRN IFHLFHDVVP TYHKENLQKL PQLAAIHHNN CMYIAHHLLT LGHQFR LRL APILCDGTAT FVDLVPGFRR LGTECFLAQM RAQKGELLER LSSARNFSNM DDEENYSAAS KAVRQVLHQL KRLGIVW QD VLPVNIYCKA MGTLLNTAIS EVIGKITALE DISTEDGDRL YSLCKTVMDE GPQVFAPLSE ESKNKKYQEE VPVYVPKW M PFKELMMMLQ ASLQEIGDRW ADGKGPLAAA FSSSEVKALI RALFQNTERR AAALAKIK

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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

Concentration5 mg/mL
BufferpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.7 µm / Nominal defocus min: 1.2 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.9 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 191979
Initial angle assignmentType: OTHER
Final angle assignmentType: OTHER
FSC plot (resolution estimation)

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