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- PDB-8q5h: Human KMN network (outer kinetochore) -

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Basic information

Entry
Database: PDB / ID: 8q5h
TitleHuman KMN network (outer kinetochore)
Components
  • (Kinetochore protein ...) x 2
  • (Kinetochore-associated protein ...) x 2
  • Kinetochore scaffold 1
  • Polyamine-modulated factor 1
  • Protein MIS12 homolog
KeywordsCELL CYCLE / OUTER KINETOCHORE / KMN / COMPLEX
Function / homology
Function and homology information


Knl1/Spc105 complex / positive regulation of meiosis I spindle assembly checkpoint / homologous chromosome orientation in meiotic metaphase I / MIS12/MIND type complex / skeletal muscle satellite cell proliferation / Ndc80 complex / leucine zipper domain binding / acrosome assembly / regulation of mitotic cell cycle spindle assembly checkpoint / attachment of spindle microtubules to kinetochore ...Knl1/Spc105 complex / positive regulation of meiosis I spindle assembly checkpoint / homologous chromosome orientation in meiotic metaphase I / MIS12/MIND type complex / skeletal muscle satellite cell proliferation / Ndc80 complex / leucine zipper domain binding / acrosome assembly / regulation of mitotic cell cycle spindle assembly checkpoint / attachment of spindle microtubules to kinetochore / outer kinetochore / attachment of mitotic spindle microtubules to kinetochore / kinetochore assembly / protein localization to kinetochore / mitotic spindle assembly checkpoint signaling / mitotic sister chromatid segregation / 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 / Resolution of Sister Chromatid Cohesion / mitotic spindle organization / acrosomal vesicle / chromosome segregation / RHO GTPases Activate Formins / kinetochore / fibrillar center / spindle pole / Separation of Sister Chromatids / azurophil granule lumen / microtubule binding / transcription regulator complex / transcription by RNA polymerase II / transcription coactivator activity / nuclear body / nuclear speck / cell division / intracellular membrane-bounded organelle / Neutrophil degranulation / nucleolus / Golgi apparatus / extracellular region / nucleoplasm / nucleus / cytosol
Similarity search - Function
Chromosome segregation protein Spc25, C-terminal / Kinetochore Mis14/Nsl1 / Kinetochore scaffold 1 / Knl1, C-terminal RWD domain / KNL1 MELT repeat / Kinetochore protein Spc25 / Chromosome segregation protein Spc25 / Kinetochore protein Mis14 like / Knl1 RWD C-terminal domain / MELT motif ...Chromosome segregation protein Spc25, C-terminal / Kinetochore Mis14/Nsl1 / Kinetochore scaffold 1 / Knl1, C-terminal RWD domain / KNL1 MELT repeat / Kinetochore protein Spc25 / Chromosome segregation protein Spc25 / Kinetochore protein Mis14 like / Knl1 RWD C-terminal domain / MELT motif / Nuclear MIS12/MIND complex subunit PMF1/Nnf1 / Centromere protein Mis12 / Nnf1 / Mis12 protein / Kinetochore-associated protein Dsn1/Mis13 / Mis12-Mtw1 protein family / Kinetochore-Ndc80 subunit Spc24 / Spc24 subunit of Ndc80
Similarity search - Domain/homology
Polyamine-modulated factor 1 / Kinetochore protein Spc24 / Kinetochore scaffold 1 / Kinetochore-associated protein NSL1 homolog / Protein MIS12 homolog / Kinetochore-associated protein DSN1 homolog / Kinetochore protein Spc25
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.5 Å
AuthorsRaisch, T. / Polley, S. / Vetter, I. / Musacchio, A. / Raunser, S.
Funding support Germany, 1items
OrganizationGrant numberCountry
Max Planck Society Germany
CitationJournal: Nat Struct Mol Biol / Year: 2024
Title: Structure of the human KMN complex and implications for regulation of its assembly.
Authors: Soumitra Polley / Tobias Raisch / Sabrina Ghetti / Marie Körner / Melina Terbeck / Frauke Gräter / Stefan Raunser / Camilo Aponte-Santamaría / Ingrid R Vetter / Andrea Musacchio /
Abstract: Biorientation of chromosomes during cell division is necessary for precise dispatching of a mother cell's chromosomes into its two daughters. Kinetochores, large layered structures built on ...Biorientation of chromosomes during cell division is necessary for precise dispatching of a mother cell's chromosomes into its two daughters. Kinetochores, large layered structures built on specialized chromosome loci named centromeres, promote biorientation by binding and sensing spindle microtubules. One of the outer layer main components is a ten-subunit assembly comprising Knl1C, Mis12C and Ndc80C (KMN) subcomplexes. The KMN is highly elongated and docks on kinetochores and microtubules through interfaces at its opposite extremes. Here, we combine cryogenic electron microscopy reconstructions and AlphaFold2 predictions to generate a model of the human KMN that reveals all intra-KMN interfaces. We identify and functionally validate two interaction interfaces that link Mis12C to Ndc80C and Knl1C. Through targeted interference experiments, we demonstrate that this mutual organization strongly stabilizes the KMN assembly. Our work thus reports a comprehensive structural and functional analysis of this part of the kinetochore microtubule-binding machinery and elucidates the path of connections from the chromatin-bound components to the force-generating components.
History
DepositionAug 9, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Feb 14, 2024Provider: repository / Type: Initial release
Revision 1.1Sep 4, 2024Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
4: Kinetochore protein Spc24
5: Kinetochore protein Spc25
A: Protein MIS12 homolog
B: Polyamine-modulated factor 1
D: Kinetochore-associated protein DSN1 homolog
K: Kinetochore scaffold 1
N: Kinetochore-associated protein NSL1 homolog


Theoretical massNumber of molelcules
Total (without water)185,9747
Polymers185,9747
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

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Kinetochore protein ... , 2 types, 2 molecules 45

#1: Protein Kinetochore protein Spc24 / hSpc24


Mass: 10344.515 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SPC24, SPBC24 / Production host: Escherichia coli (E. coli) / References: UniProt: Q8NBT2
#2: Protein Kinetochore protein Spc25 / hSpc25


Mass: 16285.332 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SPC25, SPBC25, AD024 / Production host: Escherichia coli (E. coli) / References: UniProt: Q9HBM1

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Protein , 3 types, 3 molecules ABK

#3: Protein Protein MIS12 homolog


Mass: 24170.922 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: MIS12 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9H081
#4: Protein Polyamine-modulated factor 1 / PMF-1


Mass: 23368.311 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PMF1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q6P1K2
#6: Protein Kinetochore scaffold 1 / ALL1-fused gene from chromosome 15q14 protein / AF15q14 / Bub-linking kinetochore protein / Blinkin ...ALL1-fused gene from chromosome 15q14 protein / AF15q14 / Bub-linking kinetochore protein / Blinkin / Cancer susceptibility candidate gene 5 protein / Cancer/testis antigen 29 / CT29 / Kinetochore-null protein 1 / Protein CASC5 / Protein D40/AF15q14


Mass: 38644.453 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: KNL1, CASC5, KIAA1570 / Production host: Escherichia coli (E. coli) / References: UniProt: Q8NG31

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Kinetochore-associated protein ... , 2 types, 2 molecules DN

#5: Protein Kinetochore-associated protein DSN1 homolog


Mass: 40951.062 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: DSN1, C20orf172, MIS13 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9H410
#7: Protein Kinetochore-associated protein NSL1 homolog


Mass: 32208.951 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: NSL1, C1orf48, DC31, DC8, MIS14 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q96IY1

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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: KMN network / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Trichoplusia ni (cabbage looper)
Buffer solutionpH: 8
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: OTHER
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 3000 nm / Nominal defocus min: 1500 nm
Image recordingElectron dose: 63.9 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 4.5 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 230597 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0039763
ELECTRON MICROSCOPYf_angle_d0.70713173
ELECTRON MICROSCOPYf_dihedral_angle_d4.9481259
ELECTRON MICROSCOPYf_chiral_restr0.0431452
ELECTRON MICROSCOPYf_plane_restr0.0061710

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