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- EMDB-63346: cryo-EM structure of Mis151-249-Cnp1 nucleosome complex -

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Basic information

Entry
Database: EMDB / ID: EMD-63346
Titlecryo-EM structure of Mis151-249-Cnp1 nucleosome complex
Map datacryo-EM structure of Mis151-249-Cnp1 nucleosome complex
Sample
  • Complex: cryo-EM structure of Mis151-249-Cnp1 nucleosome complex
    • Protein or peptide: Inner kinetochore subunit mis15
    • Protein or peptide: Histone H3-like centromeric protein cnp1
    • Protein or peptide: Histone H4
    • Protein or peptide: Histone H2A-beta
    • Protein or peptide: Histone H2B-alpha
    • DNA: DNA (144-MER)
    • DNA: DNA (144-MER)
KeywordsNucleosome dynamics / CENP-A / Cnp1 / kinetochore assembly / Chromosome Segregation / NUCLEAR PROTEIN/DNA / NUCLEAR PROTEIN-DNA complex
Function / homology
Function and homology information


PKMTs methylate histone lysines / HDMs demethylate histones / CENP-A containing chromatin / : / Condensation of Prophase Chromosomes / HATs acetylate histones / : / : / Assembly of the ORC complex at the origin of replication / Oxidative Stress Induced Senescence ...PKMTs methylate histone lysines / HDMs demethylate histones / CENP-A containing chromatin / : / Condensation of Prophase Chromosomes / HATs acetylate histones / : / : / Assembly of the ORC complex at the origin of replication / Oxidative Stress Induced Senescence / Factors involved in megakaryocyte development and platelet production / RMTs methylate histone arginines / Ub-specific processing proteases / HDACs deacetylate histones / chromosome, centromeric core domain / SUMOylation of chromatin organization proteins / Estrogen-dependent gene expression / RNA Polymerase I Promoter Escape / Recruitment and ATM-mediated phosphorylation of repair and signaling proteins at DNA double strand breaks / Transcriptional regulation by small RNAs / Mis6-Sim4 complex / mating-type region heterochromatin / mitotic DNA damage checkpoint signaling / chromosome, subtelomeric region / CENP-A containing chromatin assembly / centromeric DNA binding / homologous chromosome segregation / rDNA heterochromatin / condensed chromosome, centromeric region / mitotic chromosome condensation / chromatin-protein adaptor activity / mitotic G2 DNA damage checkpoint signaling / chromosome, centromeric region / pericentric heterochromatin / CENP-A containing nucleosome / chromosome segregation / double-strand break repair via homologous recombination / kinetochore / structural constituent of chromatin / heterochromatin formation / nucleosome / double-strand break repair / nucleosome assembly / site of double-strand break / chromatin remodeling / protein heterodimerization activity / cell division / chromatin / DNA binding / nucleus
Similarity search - Function
Centromere protein Chl4/mis15/CENP-N / Kinetochore protein CHL4 like / : / Histone H2B signature. / Histone H2A conserved site / Histone H2A signature. / Histone H2B / Histone H2B / Histone H2A, C-terminal domain / C-terminus of histone H2A ...Centromere protein Chl4/mis15/CENP-N / Kinetochore protein CHL4 like / : / Histone H2B signature. / Histone H2A conserved site / Histone H2A signature. / Histone H2B / Histone H2B / Histone H2A, C-terminal domain / C-terminus of histone H2A / Histone 2A / Histone H2A / Histone H4, conserved site / Histone H4 signature. / Histone H4 / Histone H4 / CENP-T/Histone H4, histone fold / Centromere kinetochore component CENP-T histone fold / Histone H3 / Histone H3/CENP-A / Histone H2A/H2B/H3 / Core histone H2A/H2B/H3/H4 domain / Histone-fold
Similarity search - Domain/homology
Histone H2A-beta / Histone H2B-alpha / Histone H4 / Inner kinetochore subunit mis15 / Histone H3-like centromeric protein cnp1
Similarity search - Component
Biological speciesSchizosaccharomyces pombe (strain 972 / ATCC 24843) (yeast) / Schizosaccharomyces pombe 972h- (yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.83 Å
AuthorsXiong Y / Zang J
Funding support1 items
OrganizationGrant numberCountry
Other government
CitationJournal: J Mol Cell Biol / Year: 2025
Title: Cnp1 N-terminal dynamics regulate L1 loop recognition by Mis15 to orchestrate kinetochore assembly in Schizosaccharomyces pombe.
Authors: Yujie Xiong / Yanze Jian / Yongliang Zhang / Min Zhang / Xuan Zhang / Kaiming Zhang / Chuanhai Fu / Tian Tian / Jianye Zang /
Abstract: Centromeres are defined by the histone H3 variant CENP-A, which serve as the foundation for kinetochore assembly and ensure faithful chromosome segregation. CENP-A nucleosomes possess distinctive ...Centromeres are defined by the histone H3 variant CENP-A, which serve as the foundation for kinetochore assembly and ensure faithful chromosome segregation. CENP-A nucleosomes possess distinctive dynamic features, including flexible DNA ends at the entry/exit sites and a mobile N-terminal region, which are properties proposed to facilitate kinetochore assembly, yet the underlying molecular mechanisms remain elusive. Here, we present cryo-electron microscopy structures of Cnp1, the Schizosaccharomyces pombe (S. pombe) ortholog of CENP-A, alone and in complex with Mis15, the fission yeast ortholog of CENP-N. By integrating structural, biochemical, and molecular dynamics analyses, we demonstrate that the N-terminal region of Cnp1 regulates both DNA-end breathing and the conformational mobility of the L1 loop, a critical structural element for Mis15 recognition. Either enhanced dynamics caused by N-terminal deletion or reduced dynamics from targeted residue substitution disrupt Mis15 binding in vitro and impair its centromeric localization in vivo, thereby compromising the earliest steps of constitutive centromere-associated network assembly. Our findings establish the Cnp1 N-terminus as a dynamic allosteric modulator of chromatin architecture and reveal an L1 loop modulation mechanism that links nucleosome flexibility to kinetochore specification and chromosome segregation fidelity in fission yeast.
History
DepositionFeb 1, 2025-
Header (metadata) releaseFeb 4, 2026-
Map releaseFeb 4, 2026-
UpdateFeb 4, 2026-
Current statusFeb 4, 2026Processing site: PDBc / Status: Released

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Structure visualization

Supplemental images

Downloads & links

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Map

FileDownload / File: emd_63346.map.gz / Format: CCP4 / Size: 144.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationcryo-EM structure of Mis151-249-Cnp1 nucleosome complex
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.07 Å/pix.
x 336 pix.
= 359.52 Å
1.07 Å/pix.
x 336 pix.
= 359.52 Å
1.07 Å/pix.
x 336 pix.
= 359.52 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.07 Å
Density
Contour LevelBy AUTHOR: 0.04
Minimum - Maximum-0.1958487 - 0.54898363
Average (Standard dev.)0.00030742283 (±0.014674948)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions336336336
Spacing336336336
CellA=B=C: 359.52002 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: cryo-EM structure of Mis151-249-Cnp1 nucleosome complex

Fileemd_63346_additional_1.map
Annotationcryo-EM structure of Mis151-249-Cnp1 nucleosome complex
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: cryo-EM structure of Mis151-249-Cnp1 nucleosome complex

Fileemd_63346_half_map_1.map
Annotationcryo-EM structure of Mis151-249-Cnp1 nucleosome complex
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: cryo-EM structure of Mis151-249-Cnp1 nucleosome complex

Fileemd_63346_half_map_2.map
Annotationcryo-EM structure of Mis151-249-Cnp1 nucleosome complex
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire : cryo-EM structure of Mis151-249-Cnp1 nucleosome complex

EntireName: cryo-EM structure of Mis151-249-Cnp1 nucleosome complex
Components
  • Complex: cryo-EM structure of Mis151-249-Cnp1 nucleosome complex
    • Protein or peptide: Inner kinetochore subunit mis15
    • Protein or peptide: Histone H3-like centromeric protein cnp1
    • Protein or peptide: Histone H4
    • Protein or peptide: Histone H2A-beta
    • Protein or peptide: Histone H2B-alpha
    • DNA: DNA (144-MER)
    • DNA: DNA (144-MER)

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Supramolecule #1: cryo-EM structure of Mis151-249-Cnp1 nucleosome complex

SupramoleculeName: cryo-EM structure of Mis151-249-Cnp1 nucleosome complex
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Schizosaccharomyces pombe (strain 972 / ATCC 24843) (yeast)

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Macromolecule #1: Inner kinetochore subunit mis15

MacromoleculeName: Inner kinetochore subunit mis15 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Schizosaccharomyces pombe 972h- (yeast)
Molecular weightTheoretical: 29.200105 KDa
Recombinant expressionOrganism: in vitro transcription vector pT7-Fluc(deltai) (others)
SequenceString: MRTELSSNIR IKHDKKIQKL LNRFPRDFLV KLCVEWIQKQ TYPPNAKDIN LEDMLDDEEW NPEAFYKNVP KSMLKRSIIH RMLVYDWPN GFYLGQIAQL EILALAHGFV SMRWTASKVH HSAEKTVLPN PLVFLELLKS ELESIFVYHT YISRHETLPI T FIRLVLWD ...String:
MRTELSSNIR IKHDKKIQKL LNRFPRDFLV KLCVEWIQKQ TYPPNAKDIN LEDMLDDEEW NPEAFYKNVP KSMLKRSIIH RMLVYDWPN GFYLGQIAQL EILALAHGFV SMRWTASKVH HSAEKTVLPN PLVFLELLKS ELESIFVYHT YISRHETLPI T FIRLVLWD SKRPTALHTY PSSKQIFYLG LMDDSDVLLH NIFLRNDVCH SLFLQCLSRL LYRLKAGSAL RPIDLVSKNL TT FCTNVGV NK

UniProtKB: Inner kinetochore subunit mis15

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Macromolecule #2: Histone H3-like centromeric protein cnp1

MacromoleculeName: Histone H3-like centromeric protein cnp1 / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Schizosaccharomyces pombe 972h- (yeast)
Molecular weightTheoretical: 13.908231 KDa
Recombinant expressionOrganism: in vitro transcription vector pT7-Fluc(deltai) (others)
SequenceString:
MAKKSLMAEP GDPIPRPRKK RYRPGTTALR EIRKYQRSTD LLIQRLPFSR IVREISSEFV ANFSTDVGLR WQSTALQCLQ EAAEAFLVH LFEDTNLCAI HAKRVTIMQR DMQLARRIRG A

UniProtKB: Histone H3-like centromeric protein cnp1

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Macromolecule #3: Histone H4

MacromoleculeName: Histone H4 / type: protein_or_peptide / ID: 3 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Schizosaccharomyces pombe 972h- (yeast)
Molecular weightTheoretical: 8.952407 KDa
Recombinant expressionOrganism: in vitro transcription vector pT7-Fluc(deltai) (others)
SequenceString:
DNIQGITKPA IRRLARRGGV KRISALVYEE TRAVLKLFLE NVIRDAVTYT EHAKRKTVTS LDVVYSLKRQ GRTIYGFGG

UniProtKB: Histone H4

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Macromolecule #4: Histone H2A-beta

MacromoleculeName: Histone H2A-beta / type: protein_or_peptide / ID: 4 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Schizosaccharomyces pombe 972h- (yeast)
Molecular weightTheoretical: 11.295081 KDa
Recombinant expressionOrganism: in vitro transcription vector pT7-Fluc(deltai) (others)
SequenceString:
SAQSRSAKAG LAFPVGRVHR LLRKGNYAQR VGAGAPVYLA AVLEYLAAEI LELAGNAARD NKKTRIIPRH LQLAIRNDEE LNKLLGHVT IAQGGVVPNI NAHLLP

UniProtKB: Histone H2A-beta

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Macromolecule #5: Histone H2B-alpha

MacromoleculeName: Histone H2B-alpha / type: protein_or_peptide / ID: 5 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Schizosaccharomyces pombe 972h- (yeast)
Molecular weightTheoretical: 10.540012 KDa
Recombinant expressionOrganism: in vitro transcription vector pT7-Fluc(deltai) (others)
SequenceString:
KNRKETYSSY IYKVLKQVHP DTGISNQAMR ILNSFVNDIF ERIATEASKL AAYNKKSTIS SREIQTAVRL ILPGELAKHA VTEGTKSVT KYSSS

UniProtKB: Histone H2B-alpha

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Macromolecule #6: DNA (144-MER)

MacromoleculeName: DNA (144-MER) / type: dna / ID: 6 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Schizosaccharomyces pombe 972h- (yeast)
Molecular weightTheoretical: 44.168133 KDa
SequenceString: (DT)(DC)(DG)(DA)(DG)(DA)(DA)(DT)(DC)(DC) (DC)(DG)(DG)(DT)(DG)(DC)(DC)(DG)(DA)(DG) (DG)(DC)(DC)(DG)(DC)(DT)(DC)(DA)(DA) (DT)(DT)(DG)(DG)(DT)(DC)(DG)(DT)(DA)(DG) (DA) (DC)(DA)(DG)(DC)(DT)(DC) ...String:
(DT)(DC)(DG)(DA)(DG)(DA)(DA)(DT)(DC)(DC) (DC)(DG)(DG)(DT)(DG)(DC)(DC)(DG)(DA)(DG) (DG)(DC)(DC)(DG)(DC)(DT)(DC)(DA)(DA) (DT)(DT)(DG)(DG)(DT)(DC)(DG)(DT)(DA)(DG) (DA) (DC)(DA)(DG)(DC)(DT)(DC)(DT)(DA) (DG)(DC)(DA)(DC)(DC)(DG)(DC)(DT)(DT)(DA) (DA)(DA) (DC)(DG)(DC)(DA)(DC)(DG)(DT) (DA)(DC)(DG)(DC)(DG)(DC)(DT)(DG)(DT)(DC) (DC)(DC)(DC) (DC)(DG)(DC)(DG)(DT)(DT) (DT)(DT)(DA)(DA)(DC)(DC)(DG)(DC)(DC)(DA) (DA)(DG)(DG)(DG) (DG)(DA)(DT)(DT)(DA) (DC)(DT)(DC)(DC)(DC)(DT)(DA)(DG)(DT)(DC) (DT)(DC)(DC)(DA)(DG) (DG)(DC)(DA)(DC) (DG)(DT)(DG)(DT)(DC)(DA)(DC)(DA)(DT)(DA) (DT)(DA)(DT)(DA)(DC)(DA) (DT)(DC)(DC) (DG)

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Macromolecule #7: DNA (144-MER)

MacromoleculeName: DNA (144-MER) / type: dna / ID: 7 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Schizosaccharomyces pombe 972h- (yeast)
Molecular weightTheoretical: 44.728477 KDa
SequenceString: (DC)(DG)(DG)(DA)(DT)(DG)(DT)(DA)(DT)(DA) (DT)(DA)(DT)(DG)(DT)(DG)(DA)(DC)(DA)(DC) (DG)(DT)(DG)(DC)(DC)(DT)(DG)(DG)(DA) (DG)(DA)(DC)(DT)(DA)(DG)(DG)(DG)(DA)(DG) (DT) (DA)(DA)(DT)(DC)(DC)(DC) ...String:
(DC)(DG)(DG)(DA)(DT)(DG)(DT)(DA)(DT)(DA) (DT)(DA)(DT)(DG)(DT)(DG)(DA)(DC)(DA)(DC) (DG)(DT)(DG)(DC)(DC)(DT)(DG)(DG)(DA) (DG)(DA)(DC)(DT)(DA)(DG)(DG)(DG)(DA)(DG) (DT) (DA)(DA)(DT)(DC)(DC)(DC)(DC)(DT) (DT)(DG)(DG)(DC)(DG)(DG)(DT)(DT)(DA)(DA) (DA)(DA) (DC)(DG)(DC)(DG)(DG)(DG)(DG) (DG)(DA)(DC)(DA)(DG)(DC)(DG)(DC)(DG)(DT) (DA)(DC)(DG) (DT)(DG)(DC)(DG)(DT)(DT) (DT)(DA)(DA)(DG)(DC)(DG)(DG)(DT)(DG)(DC) (DT)(DA)(DG)(DA) (DG)(DC)(DT)(DG)(DT) (DC)(DT)(DA)(DC)(DG)(DA)(DC)(DC)(DA)(DA) (DT)(DT)(DG)(DA)(DG) (DC)(DG)(DG)(DC) (DC)(DT)(DC)(DG)(DG)(DC)(DA)(DC)(DC)(DG) (DG)(DG)(DA)(DT)(DT)(DC) (DT)(DC)(DG) (DA)

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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

Concentration0.2 mg/mL
BufferpH: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI POLARA 300
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 55.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 4074596
CTF correctionType: NONE
Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.83 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 57310
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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