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- PDB-7sqk: Cryo-EM structure of the human augmin complex -

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Basic information

Entry
Database: PDB / ID: 7sqk
TitleCryo-EM structure of the human augmin complex
Components
  • (HAUS augmin-like complex subunit ...) x 7
  • Isoform 4 of HAUS augmin-like complex subunit 4
KeywordsCELL CYCLE / Cell Division / Mitosis / Microtubules
Function / homology
Function and homology information


HAUS complex / microtubule minus-end binding / regulation of microtubule nucleation / nuclear microtubule / mitotic spindle microtubule / microtubule nucleation / thioesterase binding / centrosome cycle / spindle assembly / Loss of Nlp from mitotic centrosomes ...HAUS complex / microtubule minus-end binding / regulation of microtubule nucleation / nuclear microtubule / mitotic spindle microtubule / microtubule nucleation / thioesterase binding / centrosome cycle / spindle assembly / Loss of Nlp from mitotic centrosomes / Loss of proteins required for interphase microtubule organization from the centrosome / Recruitment of mitotic centrosome proteins and complexes / Recruitment of NuMA to mitotic centrosomes / Anchoring of the basal body to the plasma membrane / AURKA Activation by TPX2 / mitotic spindle / microtubule cytoskeleton organization / spindle pole / Regulation of PLK1 Activity at G2/M Transition / microtubule binding / cell division / centrosome / nucleolus / plasma membrane / cytosol / cytoplasm
Similarity search - Function
HAUS augmin-like complex subunit 2, metazoa / HAUS augmin-like complex subunit 6 / HAUS augmin-like complex subunit 6, N-terminal / HAUS augmin-like complex subunit 2 / HAUS augmin-like complex subunit 7-like / HAUS augmin-like complex subunit 6 N-terminus / HAUS augmin-like complex subunit 2 / HAUS augmin-like complex subunit 4, metazoa / HAUS complex subunit 5, metazoa / HAUS augmin-like complex subunit 1 ...HAUS augmin-like complex subunit 2, metazoa / HAUS augmin-like complex subunit 6 / HAUS augmin-like complex subunit 6, N-terminal / HAUS augmin-like complex subunit 2 / HAUS augmin-like complex subunit 7-like / HAUS augmin-like complex subunit 6 N-terminus / HAUS augmin-like complex subunit 2 / HAUS augmin-like complex subunit 4, metazoa / HAUS complex subunit 5, metazoa / HAUS augmin-like complex subunit 1 / HAUS augmin-like complex subunit 5 / HAUS augmin-like complex subunit 4 / HAUS augmin-like complex subunit 4 / HAUS augmin-like complex subunit 5 / HAUS augmin-like complex subunit 3 / HAUS augmin-like complex subunit 3, N-terminal / HAUS augmin-like complex subunit 3
Similarity search - Domain/homology
HAUS augmin-like complex subunit 5 / HAUS augmin-like complex subunit 3 / HAUS augmin-like complex subunit 6 / HAUS augmin-like complex subunit 1 / HAUS augmin-like complex subunit 7 / HAUS augmin-like complex subunit 8 / HAUS augmin-like complex subunit 4 / HAUS augmin-like complex subunit 2
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 8 Å
AuthorsGabel, C.A. / Chang, L.
Funding support1items
OrganizationGrant numberCountry
Not funded
CitationJournal: Nat Commun / Year: 2022
Title: Molecular architecture of the augmin complex.
Authors: Clinton A Gabel / Zhuang Li / Andrew G DeMarco / Ziguo Zhang / Jing Yang / Mark C Hall / David Barford / Leifu Chang /
Abstract: Accurate segregation of chromosomes during mitosis depends on the correct assembly of the mitotic spindle, a bipolar structure composed mainly of microtubules. The augmin complex, or homologous to ...Accurate segregation of chromosomes during mitosis depends on the correct assembly of the mitotic spindle, a bipolar structure composed mainly of microtubules. The augmin complex, or homologous to augmin subunits (HAUS) complex, is an eight-subunit protein complex required for building robust mitotic spindles in metazoa. Augmin increases microtubule density within the spindle by recruiting the γ-tubulin ring complex (γ-TuRC) to pre-existing microtubules and nucleating branching microtubules. Here, we elucidate the molecular architecture of augmin by single particle cryo-electron microscopy (cryo-EM), computational methods, and crosslinking mass spectrometry (CLMS). Augmin's highly flexible structure contains a V-shaped head and a filamentous tail, with the head existing in either extended or contracted conformational states. Our work highlights how cryo-EM, complemented by computational advances and CLMS, can elucidate the structure of a challenging protein complex and provides insights into the function of augmin in mediating microtubule branching nucleation.
History
DepositionNov 5, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Sep 21, 2022Provider: repository / Type: Initial release
Revision 1.1Oct 5, 2022Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: HAUS augmin-like complex subunit 1
B: HAUS augmin-like complex subunit 2
C: HAUS augmin-like complex subunit 3
D: Isoform 4 of HAUS augmin-like complex subunit 4
E: HAUS augmin-like complex subunit 5
F: HAUS augmin-like complex subunit 6
G: HAUS augmin-like complex subunit 7
H: HAUS augmin-like complex subunit 8


Theoretical massNumber of molelcules
Total (without water)379,1838
Polymers379,1838
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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HAUS augmin-like complex subunit ... , 7 types, 7 molecules ABCEFGH

#1: Protein HAUS augmin-like complex subunit 1 / Coiled-coil domain-containing protein 5 / Enhancer of invasion-cluster / HEI-C


Mass: 31910.348 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: HAUS1, CCDC5, HEIC / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q96CS2
#2: Protein HAUS augmin-like complex subunit 2 / Centrosomal protein of 27 kDa / Cep27


Mass: 32487.453 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: HAUS2, C15orf25, CEP27 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9NVX0
#3: Protein HAUS augmin-like complex subunit 3


Mass: 69740.180 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: HAUS3, C4orf15 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q68CZ6
#5: Protein HAUS augmin-like complex subunit 5


Mass: 71783.406 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: HAUS5, KIAA0841 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: O94927
#6: Protein HAUS augmin-like complex subunit 6


Mass: 50250.039 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: HAUS6, DGT6, FAM29A, KIAA1574 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q7Z4H7
#7: Protein HAUS augmin-like complex subunit 7 / 26S proteasome-associated UCH37-interacting protein 1 / UCHL5-interacting protein / X-linked protein STS1769


Mass: 40819.152 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: HAUS7, UCHL5IP, UIP1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q99871
#8: Protein HAUS augmin-like complex subunit 8 / HEC1/NDC80-interacting centrosome-associated protein 1 / Sarcoma antigen NY-SAR-48


Mass: 44922.430 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: HAUS8, HICE1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9BT25

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Protein , 1 types, 1 molecules D

#4: Protein Isoform 4 of HAUS augmin-like complex subunit 4


Mass: 37269.805 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: HAUS4, C14orf94 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9H6D7

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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: Holocomplex of the Homologous to Augment Subunits Complex
Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 378 kDa/nm / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Trichoplusia ni (cabbage looper)
Buffer solutionpH: 8
Buffer component
IDConc.NameFormulaBuffer-ID
120 mM4-(2-hydroxyethyl)-1-piperazineethanesulfonic acidHEPES1
2200 mMsodium chlorideNaClSodium chloride1
31 mMtris(2-carboxyethyl)phosphineTCEP1
41 % (v/v)GlycerolGlycerol1
SpecimenConc.: 0.05 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid type: Quantifoil R2/2
VitrificationInstrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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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 FIELDBright-field microscopy / Nominal magnification: 64000 X / Nominal defocus max: 1500 nm / Nominal defocus min: 800 nm / Calibrated defocus min: 800 nm / Calibrated defocus max: 1500 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 50 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k)
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV / Phase plate: VOLTA PHASE PLATE

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Processing

SoftwareName: PHENIX / Version: 1.19.1_4122: / Classification: refinement
EM software
IDNameVersionCategory
2Leginon3.4image acquisition
4GctfCTF correction
7Coot9.05model fitting
13PHENIX1.19.1model refinement
CTF correctionType: PHASE FLIPPING ONLY
Particle selectionNum. of particles selected: 2000000
3D reconstructionResolution: 8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 447000 / Algorithm: FOURIER SPACE / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00423969
ELECTRON MICROSCOPYf_angle_d0.89332306
ELECTRON MICROSCOPYf_dihedral_angle_d5.1633132
ELECTRON MICROSCOPYf_chiral_restr0.0473663
ELECTRON MICROSCOPYf_plane_restr0.0064169

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