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- PDB-8qv2: Structure of the native y-Tubulin Ring Complex (yTuRC) capping mi... -

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Basic information

Entry
Database: PDB / ID: 8qv2
TitleStructure of the native y-Tubulin Ring Complex (yTuRC) capping microtubule minus ends at the spindle pole body
Components
  • (Spindle pole body ...) x 3
  • Tubulin alpha-1 chain
  • Tubulin beta chain
  • Tubulin gamma chain
  • Unkown protein
KeywordsCELL CYCLE / Microtubule nucleation / MTOC / y-tubulin / SPB
Function / homology
Function and homology information


Cilium Assembly / nuclear migration by microtubule mediated pushing forces / nuclear division / Sealing of the nuclear envelope (NE) by ESCRT-III / nuclear migration along microtubule / homologous chromosome segregation / gamma-tubulin complex / Platelet degranulation / microtubule nucleation / spindle pole body ...Cilium Assembly / nuclear migration by microtubule mediated pushing forces / nuclear division / Sealing of the nuclear envelope (NE) by ESCRT-III / nuclear migration along microtubule / homologous chromosome segregation / gamma-tubulin complex / Platelet degranulation / microtubule nucleation / spindle pole body / gamma-tubulin binding / tubulin complex / mitotic sister chromatid segregation / microtubule-based process / cytoplasmic microtubule organization / nuclear periphery / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / structural constituent of cytoskeleton / spindle / microtubule cytoskeleton organization / spindle pole / mitotic cell cycle / microtubule / hydrolase activity / GTPase activity / GTP binding / metal ion binding / nucleus / cytoplasm
Similarity search - Function
Spindle pole body component 110, C-terminal / Spindle pole body component 110 C-terminal domain / 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-beta mRNA autoregulation signal. ...Spindle pole body component 110, C-terminal / Spindle pole body component 110 C-terminal domain / 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-beta mRNA autoregulation signal. / Alpha tubulin / Beta tubulin, autoregulation binding site / Beta tubulin / 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 / GUANOSINE-5'-TRIPHOSPHATE / Tubulin beta chain / SPC98 isoform 1 / Tubulin gamma chain / Spindle pole body component / Spindle pole body component 110 / Tubulin alpha-1 chain
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
MethodELECTRON MICROSCOPY / subtomogram averaging / cryo EM / Resolution: 9.2 Å
AuthorsDendooven, T. / Yatskevich, S. / Burt, A. / Bellini, D. / Kilmartin, J. / Barford, D.
Funding support United Kingdom, Germany, 3items
OrganizationGrant numberCountry
Cancer Research UK United Kingdom
Boehringer Ingelheim Fonds (BIF) Germany
Medical Research Council (MRC, United Kingdom) United Kingdom
CitationJournal: Nat Struct Mol Biol / Year: 2024
Title: Structure of the native γ-tubulin ring complex capping spindle microtubules.
Authors: Tom Dendooven / Stanislau Yatskevich / Alister Burt / Zhuo A Chen / Dom Bellini / Juri Rappsilber / John V Kilmartin / David Barford /
Abstract: Microtubule (MT) filaments, composed of α/β-tubulin dimers, are fundamental to cellular architecture, function and organismal development. They are nucleated from MT organizing centers by the ...Microtubule (MT) filaments, composed of α/β-tubulin dimers, are fundamental to cellular architecture, function and organismal development. They are nucleated from MT organizing centers by the evolutionarily conserved γ-tubulin ring complex (γTuRC). However, the molecular mechanism of nucleation remains elusive. Here we used cryo-electron tomography to determine the structure of the native γTuRC capping the minus end of a MT in the context of enriched budding yeast spindles. In our structure, γTuRC presents a ring of γ-tubulin subunits to seed nucleation of exclusively 13-protofilament MTs, adopting an active closed conformation to function as a perfect geometric template for MT nucleation. Our cryo-electron tomography reconstruction revealed that a coiled-coil protein staples the first row of α/β-tubulin of the MT to alternating positions along the γ-tubulin ring of γTuRC. This positioning of α/β-tubulin onto γTuRC suggests a role for the coiled-coil protein in augmenting γTuRC-mediated MT nucleation. Based on our results, we describe a molecular model for budding yeast γTuRC activation and MT nucleation.
History
DepositionOct 17, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Apr 24, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
b: Tubulin gamma chain
c: Tubulin gamma chain
d: Tubulin gamma chain
e: Tubulin gamma chain
f: Tubulin gamma chain
g: Tubulin gamma chain
h: Tubulin gamma chain
i: Tubulin gamma chain
j: Tubulin gamma chain
k: Tubulin gamma chain
l: Tubulin gamma chain
m: Tubulin gamma chain
n: Tubulin gamma chain
C: Spindle pole body component
D: Spindle pole body component
E: Spindle pole body component
F: Spindle pole body component
G: Spindle pole body component
H: Spindle pole body component
I: Spindle pole body component
J: Spindle pole body component
K: Spindle pole body component
L: Spindle pole body component
M: Spindle pole body component
N: Spindle pole body component
O: Spindle pole body component
P: Spindle pole body component
a: Tubulin gamma chain
Ad: Tubulin alpha-1 chain
Ac: Tubulin alpha-1 chain
Bd: Tubulin beta chain
Bc: Tubulin beta chain
Af: Tubulin alpha-1 chain
Ae: Tubulin alpha-1 chain
Bf: Tubulin beta chain
Be: Tubulin beta chain
Ah: Tubulin alpha-1 chain
Ag: Tubulin alpha-1 chain
Bh: Tubulin beta chain
Bg: Tubulin beta chain
Aj: Tubulin alpha-1 chain
Ai: Tubulin alpha-1 chain
Bj: Tubulin beta chain
Bi: Tubulin beta chain
Al: Tubulin alpha-1 chain
Ak: Tubulin alpha-1 chain
Bl: Tubulin beta chain
Bk: Tubulin beta chain
An: Tubulin alpha-1 chain
Am: Tubulin alpha-1 chain
Bn: Tubulin beta chain
Bm: Tubulin beta chain
Ab: Tubulin alpha-1 chain
Bb: Tubulin beta chain
Sc: Spindle pole body component 110
Sd: Spindle pole body component 110
Se: Spindle pole body component 110
Sf: Spindle pole body component 110
Ue: Unkown protein
Uf: Unkown protein
Sg: Spindle pole body component 110
Sh: Spindle pole body component 110
Si: Spindle pole body component 110
Sj: Spindle pole body component 110
Ui: Unkown protein
Uj: Unkown protein
Sk: Spindle pole body component 110
Sl: Spindle pole body component 110
Um: Unkown protein
Un: Unkown protein
Sa: Spindle pole body component 110
Sb: Spindle pole body component 110
Ua: Unkown protein
Ub: Unkown protein
Sm: Spindle pole body component 110
Sn: Spindle pole body component 110
Ap: Tubulin alpha-1 chain
Ao: Tubulin alpha-1 chain
Bp: Tubulin beta chain
Bo: Tubulin beta chain
Ar: Tubulin alpha-1 chain
Aq: Tubulin alpha-1 chain
Br: Tubulin beta chain
Bq: Tubulin beta chain
Ug: Unkown protein
Uh: Unkown protein
Uk: Unkown protein
Ul: Unkown protein
Uc: Unkown protein
Ud: Unkown protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)5,473,442104
Polymers5,466,19790
Non-polymers7,24514
Water1267
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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Protein , 4 types, 62 molecules bcdefghijklmnaAdAcAfAeAhAgAjAiAlAkAnAmAbApAoAr...

#1: Protein
Tubulin gamma chain


Mass: 52671.188 Da / Num. of mol.: 14 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: A0A8H4BZN3
#4: Protein
Tubulin alpha-1 chain


Mass: 49853.867 Da / Num. of mol.: 17 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P09733
#5: Protein
Tubulin beta chain


Mass: 50967.457 Da / Num. of mol.: 17 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: A0A6A5PXT5
#7: Protein
Unkown protein


Mass: 5720.042 Da / Num. of mol.: 14 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast)

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Spindle pole body ... , 3 types, 28 molecules CEGIKMODFHJLNPScSdSeSfSgShSiSjSkSlSaSbSmSn

#2: Protein
Spindle pole body component /


Mass: 96940.594 Da / Num. of mol.: 7 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: A0A8H4C290
#3: Protein
Spindle pole body component /


Mass: 98336.211 Da / Num. of mol.: 7 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: A0A8H4BVY6
#6: Protein
Spindle pole body component 110 /


Mass: 111987.125 Da / Num. of mol.: 14 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: A0A8H8UNQ3

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Non-polymers , 3 types, 21 molecules

#8: Chemical
ChemComp-GTP / GUANOSINE-5'-TRIPHOSPHATE / Guanosine triphosphate


Mass: 523.180 Da / Num. of mol.: 13 / Source method: obtained synthetically / Formula: C10H16N5O14P3 / Comment: GTP, energy-carrying molecule*YM
#9: Chemical ChemComp-GDP / GUANOSINE-5'-DIPHOSPHATE / Guanosine diphosphate


Type: RNA linking / Mass: 443.201 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H15N5O11P2 / Comment: GDP, energy-carrying molecule*YM
#10: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 7 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: subtomogram averaging

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Sample preparation

ComponentName: y-Tubulin Ring Complex capping the microtubule minus end
Type: ORGANELLE OR CELLULAR COMPONENT / Entity ID: #1-#7 / Source: NATURAL
Molecular weightExperimental value: NO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Buffer solutionpH: 6.53
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: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 4500 nm / Nominal defocus min: 2000 nm / C2 aperture diameter: 50 µm
Image recordingElectron dose: 3 e/Å2 / Avg electron dose per subtomogram: 123 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM softwareName: PHENIX / Version: 1.17.1_3660: / Category: model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 9.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 7910 / Symmetry type: POINT
EM volume selectionNum. of tomograms: 364 / Num. of volumes extracted: 31720
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.005261991
ELECTRON MICROSCOPYf_angle_d0.443355044
ELECTRON MICROSCOPYf_dihedral_angle_d4.35435406
ELECTRON MICROSCOPYf_chiral_restr0.03739414
ELECTRON MICROSCOPYf_plane_restr0.00346349

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