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- PDB-7xhn: Structure of human inner kinetochore CCAN-DNA complex -

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Entry
Database: PDB / ID: 7xhn
TitleStructure of human inner kinetochore CCAN-DNA complex
Components
  • (Centromere protein ...) x 15
  • (DNA (25-MER)) x 2
  • CENP-W
KeywordsCELL CYCLE
Function / homology
Function and homology information


FANCM-MHF complex / Mis6-Sim4 complex / positive regulation of protein localization to kinetochore / : / kinetochore organization / Fanconi anaemia nuclear complex / spindle attachment to meiosis I kinetochore / metaphase chromosome alignment / kinetochore binding / CENP-A containing chromatin assembly ...FANCM-MHF complex / Mis6-Sim4 complex / positive regulation of protein localization to kinetochore / : / kinetochore organization / Fanconi anaemia nuclear complex / spindle attachment to meiosis I kinetochore / metaphase chromosome alignment / kinetochore binding / CENP-A containing chromatin assembly / sex differentiation / centromeric DNA binding / resolution of meiotic recombination intermediates / chordate embryonic development / negative regulation of epithelial cell apoptotic process / kinetochore assembly / attachment of mitotic spindle microtubules to kinetochore / condensed chromosome, centromeric region / inner kinetochore / replication fork processing / mitotic sister chromatid segregation / chromosome, centromeric region / chromosome organization / interstrand cross-link repair / pericentric heterochromatin / Amplification of signal from unattached kinetochores via a MAD2 inhibitory signal / Mitotic Prometaphase / EML4 and NUDC in mitotic spindle formation / Deposition of new CENPA-containing nucleosomes at the centromere / NRIF signals cell death from the nucleus / Resolution of Sister Chromatid Cohesion / positive regulation of protein ubiquitination / positive regulation of epithelial cell proliferation / mitotic spindle organization / chromosome segregation / Fanconi Anemia Pathway / RHO GTPases Activate Formins / PKR-mediated signaling / kinetochore / centriolar satellite / nuclear matrix / Separation of Sister Chromatids / mitotic cell cycle / actin cytoskeleton / chromosome / midbody / cell adhesion / nuclear body / protein heterodimerization activity / cell division / DNA repair / apoptotic process / DNA damage response / chromatin binding / regulation of DNA-templated transcription / chromatin / nucleolus / signal transduction / DNA binding / nucleoplasm / identical protein binding / nucleus / membrane / cytoplasm / cytosol
Similarity search - Function
Centromere protein W / : / CENP-W protein / Centromere protein T / Centromere kinetochore component CENP-T, N-terminal domain / Centromere kinetochore component CENP-T N-terminus / Centromere subunit L / Centromere protein L / Centromere protein R / Centromere protein Q ...Centromere protein W / : / CENP-W protein / Centromere protein T / Centromere kinetochore component CENP-T, N-terminal domain / Centromere kinetochore component CENP-T N-terminus / Centromere subunit L / Centromere protein L / Centromere protein R / Centromere protein Q / Centromere protein P / Kinetochore component, CENP-R / CENP-Q, a CENPA-CAD centromere complex subunit / CENP-A-nucleosome distal (CAD) centromere subunit, CENP-P / Centromere protein H, C-terminal / Centromere protein Cenp-M / Centromere protein Cenp-K / Centromere protein H / Centromere protein H (CENP-H) / Centromere protein M (CENP-M) / Centromere-associated protein K / Centromere protein I / Mis6 / Centromere protein U / CENP-A nucleosome associated complex (NAC) subunit / Centromere protein X / CENP-S/Mhf1 / CENP-S associating Centromere protein X / CENP-S protein / CENP-C, middle DNMT3B-binding domain / Centromere assembly component CENP-C middle DNMT3B-binding region / Centromere protein O / : / Cenp-O kinetochore centromere component / Kinetochore assembly subunit CENP-C, N-terminal domain / Kinetochore assembly subunit CENP-C N-terminal / Mif2/CENP-C cupin domain / Centromere protein C/Mif2/cnp3 / Mif2/CENP-C like / Centromere protein Chl4/mis15/CENP-N / Kinetochore protein CHL4 like / Histone, subunit A / Histone, subunit A / RmlC-like cupin domain superfamily / RmlC-like jelly roll fold / CENP-T/Histone H4, histone fold / Centromere kinetochore component CENP-T histone fold / Histone-fold / P-loop containing nucleotide triphosphate hydrolases / Rossmann fold / P-loop containing nucleoside triphosphate hydrolase / Orthogonal Bundle / 3-Layer(aba) Sandwich / Mainly Alpha / Alpha Beta
Similarity search - Domain/homology
DNA / DNA (> 10) / Centromere protein X / Centromere protein C / Centromere protein R / Centromere protein W / Centromere protein P / Centromere protein U / Centromere protein Q / Centromere protein L ...DNA / DNA (> 10) / Centromere protein X / Centromere protein C / Centromere protein R / Centromere protein W / Centromere protein P / Centromere protein U / Centromere protein Q / Centromere protein L / Centromere protein S / Centromere protein I / Centromere protein T / Centromere protein N / Centromere protein K / Centromere protein O / Centromere protein H / Centromere protein M
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.71 Å
AuthorsSun, L.F. / Tian, T. / Wang, C.L. / Yang, Z.S. / Zang, J.Y.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC) China
CitationJournal: Cell Discov / Year: 2022
Title: Structural insights into human CCAN complex assembled onto DNA.
Authors: Tian Tian / Lili Chen / Zhen Dou / Zhisen Yang / Xinjiao Gao / Xiao Yuan / Chengliang Wang / Ran Liu / Zuojun Shen / Ping Gui / Maikun Teng / Xianlei Meng / Donald L Hill / Lin Li / Xuan ...Authors: Tian Tian / Lili Chen / Zhen Dou / Zhisen Yang / Xinjiao Gao / Xiao Yuan / Chengliang Wang / Ran Liu / Zuojun Shen / Ping Gui / Maikun Teng / Xianlei Meng / Donald L Hill / Lin Li / Xuan Zhang / Xing Liu / Linfeng Sun / Jianye Zang / Xuebiao Yao /
Abstract: In mitosis, accurate chromosome segregation depends on kinetochores that connect centromeric chromatin to spindle microtubules. The centromeres of budding yeast, which are relatively simple, are ...In mitosis, accurate chromosome segregation depends on kinetochores that connect centromeric chromatin to spindle microtubules. The centromeres of budding yeast, which are relatively simple, are connected to individual microtubules via a kinetochore constitutive centromere associated network (CCAN). However, the complex centromeres of human chromosomes comprise millions of DNA base pairs and attach to multiple microtubules. Here, by use of cryo-electron microscopy and functional analyses, we reveal the molecular basis of how human CCAN interacts with duplex DNA and facilitates accurate chromosome segregation. The overall structure relates to the cooperative interactions and interdependency of the constituent sub-complexes of the CCAN. The duplex DNA is topologically entrapped by human CCAN. Further, CENP-N does not bind to the RG-loop of CENP-A but to DNA in the CCAN complex. The DNA binding activity is essential for CENP-LN localization to centromere and chromosome segregation during mitosis. Thus, these analyses provide new insights into mechanisms of action underlying kinetochore assembly and function in mitosis.
History
DepositionApr 9, 2022Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jan 25, 2023Provider: repository / Type: Initial release
Revision 1.1Jul 3, 2024Group: Data collection / Category: chem_comp_atom / chem_comp_bond / em_admin / Item: _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
o: Centromere protein O
H: Centromere protein H
I: Centromere protein I
K: Centromere protein K
L: Centromere protein L
M: Centromere protein M
N: Centromere protein N
O: Centromere protein O
P: Centromere protein P
S: Centromere protein S
T: Centromere protein T
W: CENP-W
X: Centromere protein X
G: DNA (25-MER)
J: DNA (25-MER)
C: Centromere protein C
c: Centromere protein C
Q: Centromere protein Q
U: Centromere protein U
R: Centromere protein R


Theoretical massNumber of molelcules
Total (without water)772,18120
Polymers772,18120
Non-polymers00
Water00
1


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

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Components

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Centromere protein ... , 15 types, 17 molecules oOHIKLMNPSTXCcQUR

#1: Protein Centromere protein O / CENP-O / Interphase centromere complex protein 36


Mass: 33830.637 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CENPO / Production host: Trichoplusia ni (cabbage looper) / Strain (production host): Hi5 / References: UniProt: Q9BU64
#2: Protein Centromere protein H / CENP-H / Interphase centromere complex protein 35


Mass: 29349.811 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CENPH / Production host: Trichoplusia ni (cabbage looper) / Strain (production host): Hi5 / References: UniProt: Q9H3R5
#3: Protein Centromere protein I / CENP-I / FSH primary response protein 1 / Follicle-stimulating hormone primary response protein / ...CENP-I / FSH primary response protein 1 / Follicle-stimulating hormone primary response protein / Interphase centromere complex protein 19 / Leucine-rich primary response protein 1


Mass: 86820.188 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CENPI / Production host: Trichoplusia ni (cabbage looper) / Strain (production host): Hi5 / References: UniProt: Q92674
#4: Protein Centromere protein K / CENP-K / Interphase centromere complex protein 37 / Protein AF-5alpha / p33


Mass: 31696.070 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CENPK / Production host: Trichoplusia ni (cabbage looper) / Strain (production host): Hi5 / References: UniProt: Q9BS16
#5: Protein Centromere protein L / CENP-L / Interphase centromere complex protein 33


Mass: 39039.641 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CENPL / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q8N0S6
#6: Protein Centromere protein M / CENP-M / Interphase centromere complex protein 39 / Proliferation-associated nuclear element protein 1


Mass: 19761.945 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CENPM / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9NSP4
#7: Protein Centromere protein N / CENP-N / Interphase centromere complex protein 32


Mass: 40438.418 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CENPN / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q96H22
#8: Protein Centromere protein P / CENP-P


Mass: 33210.949 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CENPP / Production host: Trichoplusia ni (cabbage looper) / Strain (production host): Hi5 / References: UniProt: Q6IPU0
#9: Protein Centromere protein S / CENP-S / Apoptosis-inducing TAF9-like domain-containing protein 1 / FANCM-associated histone fold ...CENP-S / Apoptosis-inducing TAF9-like domain-containing protein 1 / FANCM-associated histone fold protein 1 / FANCM-interacting histone fold protein 1 / Fanconi anemia-associated polypeptide of 16 kDa


Mass: 15917.875 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CENPS / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q8N2Z9
#10: Protein Centromere protein T / CENP-T / Interphase centromere complex protein 22


Mass: 60502.613 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CENPT / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q96BT3
#12: Protein Centromere protein X / CENP-X / FANCM-associated histone fold protein 2 / FANCM-interacting histone fold protein 2 / ...CENP-X / FANCM-associated histone fold protein 2 / FANCM-interacting histone fold protein 2 / Fanconi anemia-associated polypeptide of 10 kDa / Retinoic acid-inducible gene D9 protein homolog / Stimulated by retinoic acid gene 13 protein homolog


Mass: 8972.415 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CENPX / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: A8MT69
#15: Protein Centromere protein C / CENP-C / Centromere autoantigen C / Centromere protein C 1 / CENP-C 1 / Interphase centromere ...CENP-C / Centromere autoantigen C / Centromere protein C 1 / CENP-C 1 / Interphase centromere complex protein 7


Mass: 107022.273 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CENPC / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q03188
#16: Protein Centromere protein Q / CENP-Q


Mass: 31477.244 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CENPQ / Production host: Trichoplusia ni (cabbage looper) / Strain (production host): Hi5 / References: UniProt: Q7L2Z9
#17: Protein Centromere protein U / CENP-U / Centromere protein of 50 kDa / CENP-50 / Interphase centromere complex protein 24 / KSHV ...CENP-U / Centromere protein of 50 kDa / CENP-50 / Interphase centromere complex protein 24 / KSHV latent nuclear antigen-interacting protein 1 / MLF1-interacting protein / Polo-box-interacting protein 1


Mass: 47609.766 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CENPU / Production host: Trichoplusia ni (cabbage looper) / Strain (production host): Hi5 / References: UniProt: Q71F23
#18: Protein Centromere protein R / CENP-R / Beta-3-endonexin / Integrin beta-3-binding protein / Nuclear receptor-interacting factor 3


Mass: 20228.297 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CENPR / Production host: Trichoplusia ni (cabbage looper) / Strain (production host): Hi5 / References: UniProt: Q13352

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Protein , 1 types, 1 molecules W

#11: Protein CENP-W


Mass: 10087.236 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q5EE01

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DNA chain , 2 types, 2 molecules GJ

#13: DNA chain DNA (25-MER)


Mass: 7685.937 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Homo sapiens (human)
#14: DNA chain DNA (25-MER)


Mass: 7676.923 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Homo sapiens (human)

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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: CCAN-DNA / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm)
Buffer solutionpH: 7.5
SpecimenEmbedding 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: SPOT SCAN
Electron lensMode: DIFFRACTION / Nominal defocus max: 2700 nm / Nominal defocus min: 1700 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K2 QUANTUM (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.20.1_4487: / Classification: refinement
CTF correctionType: NONE
3D reconstructionResolution: 3.71 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 79777 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.01123862
ELECTRON MICROSCOPYf_angle_d1.19332474
ELECTRON MICROSCOPYf_dihedral_angle_d14.6483540
ELECTRON MICROSCOPYf_chiral_restr0.0573802
ELECTRON MICROSCOPYf_plane_restr0.0073937

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