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- PDB-9h9p: Spokes 12 and 13 of the human gamma-tubulin ring complex in compl... -

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Basic information

Entry
Database: PDB / ID: 9h9p
TitleSpokes 12 and 13 of the human gamma-tubulin ring complex in complex with CDK5RAP2 and docked MZT2/GCP2-NHD module
Components
  • CDK5 regulatory subunit-associated protein 2
  • Gamma-tubulin complex component 6
  • Isoform 3 of Gamma-tubulin complex component 2
  • Mitotic-spindle organizing protein 2A
  • Tubulin gamma-1 chain
KeywordsCELL CYCLE / cytoskeleton / microtubule / microtubule nucleation / complex / template / cap / gamma-tubulin / gamma-tubulin ring complex / CDK5RAP2 / Homo / sapiens / GCP6 / GCP2 / gamma-tubulin complex protein / MZT2 / Mozart / Mozart2 / module / CM1 / CM1 motif / CEP215
Function / homology
Function and homology information


microtubule nucleator activity / negative regulation of centriole replication / regulation of mitotic cell cycle spindle assembly checkpoint / microtubule organizing center organization / polar microtubule / microtubule plus-end / gamma-tubulin complex / gamma-tubulin ring complex / mitotic spindle microtubule / meiotic spindle organization ...microtubule nucleator activity / negative regulation of centriole replication / regulation of mitotic cell cycle spindle assembly checkpoint / microtubule organizing center organization / polar microtubule / microtubule plus-end / gamma-tubulin complex / gamma-tubulin ring complex / mitotic spindle microtubule / meiotic spindle organization / microtubule nucleation / microtubule bundle formation / gamma-tubulin binding / non-motile cilium / centrosome cycle / regulation of neuron differentiation / pericentriolar material / mitotic spindle pole / cell leading edge / microtubule organizing center / centriole replication / establishment of mitotic spindle orientation / mitotic sister chromatid segregation / negative regulation of neuron differentiation / cytoplasmic microtubule / spindle assembly / neurogenesis / cytoplasmic microtubule organization / positive regulation of microtubule polymerization / centriole / Loss of Nlp from mitotic centrosomes / Loss of proteins required for interphase microtubule organization from the centrosome / Recruitment of mitotic centrosome proteins and complexes / tubulin binding / Recruitment of NuMA to mitotic centrosomes / Anchoring of the basal body to the plasma membrane / AURKA Activation by TPX2 / condensed nuclear chromosome / mitotic spindle organization / meiotic cell cycle / chromosome segregation / brain development / recycling endosome / structural constituent of cytoskeleton / microtubule cytoskeleton organization / spindle / neuron migration / spindle pole / cell junction / apical part of cell / Regulation of PLK1 Activity at G2/M Transition / mitotic cell cycle / protein-containing complex assembly / microtubule binding / microtubule / cytoskeleton / transcription cis-regulatory region binding / calmodulin binding / neuron projection / ciliary basal body / centrosome / protein-containing complex binding / protein kinase binding / GTP binding / positive regulation of DNA-templated transcription / perinuclear region of cytoplasm / Golgi apparatus / extracellular exosome / nucleoplasm / identical protein binding / nucleus / membrane / cytosol / cytoplasm
Similarity search - Function
CDK5 regulatory subunit-associated protein 2 / : / CDK5 regulatory subunit-associated protein 2-like, coiled coil / MOZART2 family / Mitotic-spindle organizing gamma-tubulin ring associated / Centrosomin, N-terminal motif 1 / Centrosomin N-terminal motif 1 / Gamma-tubulin complex component 6, N-terminal / Gamma-tubulin complex component 6 N-terminus / Gamma tubulin ...CDK5 regulatory subunit-associated protein 2 / : / CDK5 regulatory subunit-associated protein 2-like, coiled coil / MOZART2 family / Mitotic-spindle organizing gamma-tubulin ring associated / Centrosomin, N-terminal motif 1 / Centrosomin N-terminal motif 1 / Gamma-tubulin complex component 6, N-terminal / Gamma-tubulin complex component 6 N-terminus / Gamma tubulin / Gamma tubulin complex component, C-terminal / Gamma-tubulin complex component, C-terminal domain superfamily / Gamma tubulin complex component C-terminal / Gamma-tubulin complex component protein / Gamma tubulin complex component protein, N-terminal / Gamma tubulin complex component N-terminal / Tubulin / Tubulin, C-terminal / Tubulin C-terminal domain / Tubulin, conserved site / Tubulin subunits alpha, beta, and gamma signature. / Tubulin/FtsZ family, C-terminal domain / Tubulin/FtsZ-like, C-terminal domain / Tubulin/FtsZ, C-terminal / Tubulin/FtsZ, 2-layer sandwich domain / Tubulin/FtsZ family, GTPase domain / Tubulin/FtsZ family, GTPase domain / Tubulin/FtsZ, GTPase domain / Tubulin/FtsZ, GTPase domain superfamily
Similarity search - Domain/homology
GUANOSINE-5'-DIPHOSPHATE / Tubulin gamma-1 chain / Mitotic-spindle organizing protein 2A / Gamma-tubulin complex component 6 / CDK5 regulatory subunit-associated protein 2 / Gamma-tubulin complex component 2
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.5 Å
AuthorsVermeulen, B.J.A. / Pfeffer, S.
Funding support Germany, 6items
OrganizationGrant numberCountry
German Research Foundation (DFG)Schi 295/4-4 Germany
German Research Foundation (DFG)PF 963/1-4 Germany
German Research Foundation (DFG)PF 963/4-1 Germany
Other private
German Research Foundation (DFG)INST 35/1314-1 FUGG Germany
German Research Foundation (DFG)INST 35/1134-1 FUGG Germany
Citation
Journal: Nat Commun / Year: 2025
Title: Structural insights into the interplay between microtubule polymerases, γ-tubulin complexes and their receptors.
Authors: Anjun Zheng / Bram J A Vermeulen / Martin Würtz / Annett Neuner / Nicole Lübbehusen / Matthias P Mayer / Elmar Schiebel / Stefan Pfeffer /
Abstract: The γ-tubulin ring complex (γ-TuRC) is a structural template for controlled nucleation of microtubules from α/β-tubulin heterodimers. At the cytoplasmic side of the yeast spindle pole body, the ...The γ-tubulin ring complex (γ-TuRC) is a structural template for controlled nucleation of microtubules from α/β-tubulin heterodimers. At the cytoplasmic side of the yeast spindle pole body, the CM1-containing receptor protein Spc72 promotes γ-TuRC assembly from seven γ-tubulin small complexes (γ-TuSCs) and recruits the microtubule polymerase Stu2, yet their molecular interplay remains unclear. Here, we determine the cryo-EM structure of the Candida albicans cytoplasmic nucleation unit at 3.6 Å resolution, revealing how the γ-TuRC is assembled and conformationally primed for microtubule nucleation by the dimerised Spc72 CM1 motif. Two coiled-coil regions of Spc72 interact with the conserved C-terminal α-helix of Stu2 and thereby position the α/β-tubulin-binding TOG domains of Stu2 in the vicinity of the microtubule assembly site. Collectively, we reveal the function of CM1 motifs in γ-TuSC oligomerisation and the recruitment of microtubule polymerases to the γ-TuRC.
#1: Journal: Cell Rep / Year: 2020
Title: MZT Proteins Form Multi-Faceted Structural Modules in the γ-Tubulin Ring Complex.
Authors: Michal Wieczorek / Tzu-Lun Huang / Linas Urnavicius / Kuo-Chiang Hsia / Tarun M Kapoor /
Abstract: Microtubule organization depends on the γ-tubulin ring complex (γ-TuRC), a ∼2.3-MDa nucleation factor comprising an asymmetric assembly of γ-tubulin and GCP2-GCP6. However, it is currently ...Microtubule organization depends on the γ-tubulin ring complex (γ-TuRC), a ∼2.3-MDa nucleation factor comprising an asymmetric assembly of γ-tubulin and GCP2-GCP6. However, it is currently unclear how the γ-TuRC-associated microproteins MZT1 and MZT2 contribute to the structure and regulation of the holocomplex. Here, we report cryo-EM structures of MZT1 and MZT2 in the context of the native human γ-TuRC. MZT1 forms two subcomplexes with the N-terminal α-helical domains of GCP3 or GCP6 (GCP-NHDs) within the γ-TuRC "lumenal bridge." We determine the X-ray structure of recombinant MZT1/GCP6-NHD and find it is similar to that within the native γ-TuRC. We identify two additional MZT/GCP-NHD-like subcomplexes, one of which is located on the outer face of the γ-TuRC and comprises MZT2 and GCP2-NHD in complex with a centrosomin motif 1 (CM1)-containing peptide. Our data reveal how MZT1 and MZT2 establish multi-faceted, structurally mimetic "modules" that can expand structural and regulatory interfaces in the γ-TuRC.
History
DepositionOct 31, 2024Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jan 15, 2025Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: CDK5 regulatory subunit-associated protein 2
G: CDK5 regulatory subunit-associated protein 2
L: Gamma-tubulin complex component 6
M: Isoform 3 of Gamma-tubulin complex component 2
l: Tubulin gamma-1 chain
m: Tubulin gamma-1 chain
C: Mitotic-spindle organizing protein 2A
hetero molecules


Theoretical massNumber of molelcules
Total (without water)856,9739
Polymers856,0867
Non-polymers8862
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Protein , 5 types, 7 molecules AGLMlmC

#1: Protein CDK5 regulatory subunit-associated protein 2 / CDK5 activator-binding protein C48 / Centrosome-associated protein 215


Mass: 215417.359 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CDK5RAP2, CEP215, KIAA1633 / Production host: Escherichia coli (E. coli) / References: UniProt: Q96SN8
#2: Protein Gamma-tubulin complex component 6 / GCP-6


Mass: 200733.641 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / Cell line: Hela S3 / References: UniProt: Q96RT7
#3: Protein Isoform 3 of Gamma-tubulin complex component 2 / GCP-2 / hGCP2 / Gamma-ring complex protein 103 kDa / h103p / hGrip103 / Spindle pole body protein ...GCP-2 / hGCP2 / Gamma-ring complex protein 103 kDa / h103p / hGrip103 / Spindle pole body protein Spc97 homolog / hSpc97


Mass: 105765.719 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / Cell line: Hela S3 / References: UniProt: Q9BSJ2
#4: Protein Tubulin gamma-1 chain / Gamma-1-tubulin / Gamma-tubulin complex component 1 / GCP-1


Mass: 51255.824 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / Cell line: Hela S3 / References: UniProt: P23258
#5: Protein Mitotic-spindle organizing protein 2A / Mitotic-spindle organizing protein associated with a ring of gamma-tubulin 2A


Mass: 16240.576 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / Cell line: Hela S3 / References: UniProt: Q6P582

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Non-polymers , 1 types, 2 molecules

#6: Chemical ChemComp-GDP / GUANOSINE-5'-DIPHOSPHATE


Type: RNA linking / Mass: 443.201 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C10H15N5O11P2 / Comment: GDP, energy-carrying molecule*YM

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Details

Has ligand of interestN
Has protein modificationN

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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: Spokes 12 and 13 of the human gamma-tubulin ring complex in complex with CDK5RAP2 and docked MZT2/GCP2-NHD module
Type: COMPLEX / Entity ID: #1-#5 / Source: NATURAL
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
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: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 3500 nm / Nominal defocus min: 1500 nm
Image recordingElectron dose: 45 e/Å2 / Film or detector model: GATAN K3 (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: 137513 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 40.95 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.001916666
ELECTRON MICROSCOPYf_angle_d0.498122500
ELECTRON MICROSCOPYf_chiral_restr0.03732563
ELECTRON MICROSCOPYf_plane_restr0.00352812
ELECTRON MICROSCOPYf_dihedral_angle_d3.41842186

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