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Entry
Database: PDB / ID: 9qvn
TitleCryo-EM reconstruction of the NEDD1 anchor protein bound to the gamma-tubulin ring complex
Components
  • (Gamma-tubulin complex component ...) x 3
  • Actin, cytoplasmic 1, N-terminally processed
  • Isoform 3 of Gamma-tubulin complex component 2
  • Mitotic-spindle organizing protein 1
  • Protein NEDD1
  • TUBGCP6 protein
  • Tubulin gamma-1 chain
KeywordsSTRUCTURAL PROTEIN / Tubulin complex
Function / homology
Function and homology information


microtubule nucleation by interphase microtubule organizing center / gamma-tubulin complex localization / microtubule nucleator activity / positive regulation of norepinephrine uptake / polar microtubule / interphase microtubule organizing center / gamma-tubulin complex / gamma-tubulin ring complex / cellular response to cytochalasin B / bBAF complex ...microtubule nucleation by interphase microtubule organizing center / gamma-tubulin complex localization / microtubule nucleator activity / positive regulation of norepinephrine uptake / polar microtubule / interphase microtubule organizing center / gamma-tubulin complex / gamma-tubulin ring complex / cellular response to cytochalasin B / bBAF complex / mitotic spindle microtubule / npBAF complex / nBAF complex / brahma complex / meiotic spindle organization / regulation of transepithelial transport / Formation of annular gap junctions / morphogenesis of a polarized epithelium / Formation of the dystrophin-glycoprotein complex (DGC) / structural constituent of postsynaptic actin cytoskeleton / Gap junction degradation / GBAF complex / Folding of actin by CCT/TriC / protein localization to adherens junction / regulation of G0 to G1 transition / Cell-extracellular matrix interactions / microtubule nucleation / dense body / Tat protein binding / postsynaptic actin cytoskeleton / gamma-tubulin binding / Prefoldin mediated transfer of substrate to CCT/TriC / RSC-type complex / regulation of double-strand break repair / regulation of nucleotide-excision repair / non-motile cilium / Adherens junctions interactions / RHOF GTPase cycle / adherens junction assembly / apical protein localization / Sensory processing of sound by outer hair cells of the cochlea / Interaction between L1 and Ankyrins / tight junction / regulation of mitotic metaphase/anaphase transition / SWI/SNF complex / Sensory processing of sound by inner hair cells of the cochlea / positive regulation of T cell differentiation / apical junction complex / positive regulation of double-strand break repair / maintenance of blood-brain barrier / regulation of norepinephrine uptake / nitric-oxide synthase binding / transporter regulator activity / cortical cytoskeleton / NuA4 histone acetyltransferase complex / establishment or maintenance of cell polarity / pericentriolar material / positive regulation of stem cell population maintenance / cell leading edge / Recycling pathway of L1 / Regulation of MITF-M-dependent genes involved in pigmentation / microtubule organizing center / brush border / mitotic sister chromatid segregation / regulation of G1/S transition of mitotic cell cycle / EPH-ephrin mediated repulsion of cells / negative regulation of cell differentiation / mitotic spindle assembly / kinesin binding / RHO GTPases Activate WASPs and WAVEs / regulation of synaptic vesicle endocytosis / positive regulation of myoblast differentiation / single fertilization / RHO GTPases activate IQGAPs / regulation of protein localization to plasma membrane / positive regulation of double-strand break repair via homologous recombination / spindle assembly / cytoplasmic microtubule / cytoplasmic microtubule organization / cytoskeleton organization / EPHB-mediated forward signaling / Loss of Nlp from mitotic centrosomes / Loss of proteins required for interphase microtubule organization from the centrosome / centriole / Recruitment of mitotic centrosome proteins and complexes / substantia nigra development / Recruitment of NuMA to mitotic centrosomes / axonogenesis / Anchoring of the basal body to the plasma membrane / calyx of Held / AURKA Activation by TPX2 / nitric-oxide synthase regulator activity / condensed nuclear chromosome / mitotic spindle organization / meiotic cell cycle / Translocation of SLC2A4 (GLUT4) to the plasma membrane / FCGR3A-mediated phagocytosis / actin filament / adherens junction / positive regulation of cell differentiation
Similarity search - Function
: / Mitotic-spindle organizing protein 1 / Mitotic-spindle organizing gamma-tubulin ring associated / 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 ...: / Mitotic-spindle organizing protein 1 / Mitotic-spindle organizing gamma-tubulin ring associated / 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 / Actins signature 1. / Actin, conserved site / Actins signature 2. / Actin/actin-like conserved site / Actins and actin-related proteins signature. / Tubulin / Tubulin, C-terminal / Tubulin C-terminal domain / Tubulin, conserved site / Tubulin subunits alpha, beta, and gamma signature. / Actin / Actin family / Actin / 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 / ATPase, nucleotide binding domain / WD domain, G-beta repeat / WD40 repeat, conserved site / Trp-Asp (WD) repeats signature. / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD40 repeats / WD40 repeat / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily
Similarity search - Domain/homology
Gamma-tubulin complex component 6 / Tubulin gamma-1 chain / Actin, cytoplasmic 1 / Mitotic-spindle organizing protein 1 / Protein NEDD1 / Gamma-tubulin complex component 3 / Gamma-tubulin complex component 5 / Gamma-tubulin complex component 2 / Gamma-tubulin complex component 4
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.7 Å
AuthorsMunoz-Hernandez, H. / Xu, Y. / Wieczorek, M.
Funding support Switzerland, 2items
OrganizationGrant numberCountry
Swiss National Science FoundationTMSGI3_211309 Switzerland
Swiss National Science Foundation310030_208120 Switzerland
Citation
Journal: J Cell Biol / Year: 2025
Title: Structure of the microtubule-anchoring factor NEDD1 bound to the γ-tubulin ring complex.
Authors: Hugo Muñoz-Hernández / Yixin Xu / Aitor Pellicer Camardiel / Daniel Zhang / Allen Xue / Amol Aher / Ellie Walker / Florina Marxer / Tarun M Kapoor / Michal Wieczorek /
Abstract: The γ-tubulin ring complex (γ-TuRC) is an essential multiprotein assembly that provides a template for microtubule nucleation. The γ-TuRC is recruited to microtubule-organizing centers (MTOCs) by ...The γ-tubulin ring complex (γ-TuRC) is an essential multiprotein assembly that provides a template for microtubule nucleation. The γ-TuRC is recruited to microtubule-organizing centers (MTOCs) by the evolutionarily conserved attachment factor NEDD1. However, the structural basis of the NEDD1-γ-TuRC interaction is not known. Here, we report cryo-EM structures of NEDD1 bound to the human γ-TuRC in the absence or presence of the activating factor CDK5RAP2. We found that the C-terminus of NEDD1 forms a tetrameric α-helical assembly that contacts the lumen of the γ-TuRC cone and orients its microtubule-binding domain away from the complex. The structure of the γ-TuRC simultaneously bound to NEDD1 and CDK5RAP2 reveals that both factors can associate with the "open" conformation of the complex. Our results show that NEDD1 does not induce substantial conformational changes in the γ-TuRC but suggest that anchoring of γ-TuRC-capped microtubules by NEDD1 would be structurally compatible with the significant conformational changes experienced by the γ-TuRC during microtubule nucleation.
#1: Journal: bioRxiv / Year: 2024
Title: Structure of the microtubule anchoring factor NEDD1 bound to the γ-tubulin ring complex.
Authors: Hugo Muñoz-Hernández / Yixin Xu / Daniel Zhang / Allen Xue / Amol Aher / Aitor Pellicer Camardiel / Ellie Walker / Florina Marxer / Tarun M Kapoor / Michal Wieczorek /
Abstract: The γ-tubulin ring complex (γ-TuRC) is an essential multiprotein assembly, in which γ-tubulin, GCP2-6, actin, MZT1 and MZT2 form an asymmetric cone-shaped structure that provides a template for ...The γ-tubulin ring complex (γ-TuRC) is an essential multiprotein assembly, in which γ-tubulin, GCP2-6, actin, MZT1 and MZT2 form an asymmetric cone-shaped structure that provides a template for microtubule nucleation. The γ-TuRC is recruited to microtubule organizing centers (MTOCs), such as centrosomes and pre-existing mitotic spindle microtubules, via the evolutionarily-conserved attachment factor NEDD1. NEDD1 contains an N-terminal WD40 domain that binds to microtubules, and a C-terminal domain that associates with the γ-TuRC. However, the structural basis of the NEDD1-γ-TuRC interaction is not known. Here, we report cryo-electron microscopy (cryo-EM) structures of NEDD1 bound to the human γ-TuRC in the absence or presence of the activating factor CDK5RAP2, which interacts with GCP2 to induce conformational changes in the γ-TuRC and promote its microtubule nucleating function. We found that the C-terminus of NEDD1 forms a tetrameric α-helical assembly that contacts the lumen of the γ-TuRC cone, is anchored to GCP4, 5 and 6 via protein modules consisting of MZT1 & GCP3 subcomplexes, and orients its microtubule-binding WD40 domains away from the complex. We biochemically tested our structural models by identifying NEDD1 mutants unable to pull-down -tubulin from cultured cells. The structure of the γ-TuRC simultaneously bound to NEDD1 and CDK5RAP2 reveals that both factors can associate with the "open" conformation of the complex. Our results show that NEDD1 does not induce conformational changes in the γ-TuRC, but suggest that anchoring of γ-TuRC-capped microtubules by NEDD1 would be structurally compatible with the significant conformational changes experienced by the γ-TuRC during microtubule nucleation.
History
DepositionApr 11, 2025Deposition site: PDBE / Processing site: PDBE
Revision 1.0May 21, 2025Provider: repository / Type: Initial release
Revision 1.1Jun 4, 2025Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID / _citation_author.name / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
D: Gamma-tubulin complex component 3
F: Gamma-tubulin complex component 3
H: Gamma-tubulin complex component 3
P: Mitotic-spindle organizing protein 1
R: Mitotic-spindle organizing protein 1
S: Mitotic-spindle organizing protein 1
T: Mitotic-spindle organizing protein 1
U: Mitotic-spindle organizing protein 1
W: Protein NEDD1
X: Protein NEDD1
Z: Actin, cytoplasmic 1, N-terminally processed
a: Tubulin gamma-1 chain
b: Tubulin gamma-1 chain
c: Tubulin gamma-1 chain
e: Tubulin gamma-1 chain
f: Tubulin gamma-1 chain
g: Tubulin gamma-1 chain
h: Tubulin gamma-1 chain
i: Tubulin gamma-1 chain
j: Tubulin gamma-1 chain
k: Tubulin gamma-1 chain
l: Tubulin gamma-1 chain
m: Tubulin gamma-1 chain
n: Tubulin gamma-1 chain
r: Gamma-tubulin complex component 3
s: Gamma-tubulin complex component 3
t: Gamma-tubulin complex component 3
u: Gamma-tubulin complex component 3
v: Gamma-tubulin complex component 3
B: Gamma-tubulin complex component 3
K: Gamma-tubulin complex component 4
N: Gamma-tubulin complex component 3
d: Tubulin gamma-1 chain
V: Protein NEDD1
Y: Protein NEDD1
G: Isoform 3 of Gamma-tubulin complex component 2
L: TUBGCP6 protein
M: Isoform 3 of Gamma-tubulin complex component 2
C: Isoform 3 of Gamma-tubulin complex component 2
J: Gamma-tubulin complex component 5
A: Isoform 3 of Gamma-tubulin complex component 2
E: Isoform 3 of Gamma-tubulin complex component 2
I: Gamma-tubulin complex component 4
O: Mitotic-spindle organizing protein 1
Q: Mitotic-spindle organizing protein 1


Theoretical massNumber of molelcules
Total (without water)3,154,19345
Polymers3,154,19345
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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Gamma-tubulin complex component ... , 3 types, 13 molecules DFHrstuvBNKIJ

#1: Protein
Gamma-tubulin complex component 3 / hGCP3 / Gamma-ring complex protein 104 kDa / hGrip104 / Spindle pole body protein Spc98 homolog / hSpc98


Mass: 103710.102 Da / Num. of mol.: 10
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: TUBGCP3, GCP3 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q96CW5
#6: Protein Gamma-tubulin complex component 4 / hGCP4 / Gamma-ring complex protein 76 kDa / hGrip76


Mass: 76179.969 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: TUBGCP4, 76P, GCP4 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9UGJ1
#9: Protein Gamma-tubulin complex component 5 / GCP-5


Mass: 118467.547 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: TUBGCP5, GCP5, KIAA1899 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q96RT8

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Protein , 6 types, 32 molecules PRSTUOQWXVYZabcefghijklmndGMCAEL

#2: Protein
Mitotic-spindle organizing protein 1 / Mitotic-spindle organizing protein associated with a ring of gamma-tubulin 1


Mass: 8485.724 Da / Num. of mol.: 7
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: MZT1, C13orf37, MOZART1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q08AG7
#3: Protein
Protein NEDD1 / Neural precursor cell expressed developmentally down-regulated protein 1 / NEDD-1


Mass: 72050.984 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: NEDD1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q8NHV4
#4: Protein Actin, cytoplasmic 1, N-terminally processed


Mass: 41782.660 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ACTB / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P60709
#5: Protein
Tubulin gamma-1 chain / Gamma-1-tubulin / Gamma-tubulin complex component 1 / GCP-1


Mass: 52022.617 Da / Num. of mol.: 14
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: TUBG1, TUBG / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P23258
#7: 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.: 5
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: TUBGCP2, GCP2 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9BSJ2
#8: Protein TUBGCP6 protein


Mass: 199732.516 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: TUBGCP6 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: B2RWN4

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Details

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: cryo-EM reconstruction of the NEDD1 anchor protein bound to the gamma-tubulin ring complex
Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Spodoptera (butterflies/moths)
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: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: SPOT SCAN
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 3500 nm / Nominal defocus min: 1500 nm
Image recordingElectron dose: 60 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

EM software
IDNameVersionCategory
2EPUimage acquisition
7Cootmodel fitting
10cryoSPARCinitial Euler assignment
11cryoSPARCfinal Euler assignment
12cryoSPARCclassification
13cryoSPARC3D reconstruction
14PHENIX1.20.1_4487model refinement
CTF correctionType: NONE
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 4.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 266675 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Atomic model buildingSource name: AlphaFold / Type: in silico model
RefinementHighest resolution: 4.7 Å
Stereochemistry target values: REAL-SPACE (WEIGHTED MAP SUM AT ATOM CENTERS)
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00287683
ELECTRON MICROSCOPYf_angle_d0.512122133
ELECTRON MICROSCOPYf_dihedral_angle_d4.63217647
ELECTRON MICROSCOPYf_chiral_restr0.03816876
ELECTRON MICROSCOPYf_plane_restr0.00317647

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