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- PDB-8afy: Subtomogram average of membrane-bound Atg18 oligomers -

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Basic information

Entry
Database: PDB / ID: 8afy
TitleSubtomogram average of membrane-bound Atg18 oligomers
ComponentsAutophagy-related protein 18
KeywordsMEMBRANE PROTEIN / autophagy / membrane remodeling / PIP binding / PI3P / PI(3 / 5)P2 / lipid binding protein
Function / homology
Function and homology information


regulation of phosphatidylinositol biosynthetic process / PAS complex / 1-phosphatidyl-1D-myo-inositol 3,5-bisphosphate metabolic process / phagophore / vacuolar protein processing / positive regulation of vacuole organization / Macroautophagy / cytoplasm to vacuole targeting by the Cvt pathway / nucleophagy / pexophagy ...regulation of phosphatidylinositol biosynthetic process / PAS complex / 1-phosphatidyl-1D-myo-inositol 3,5-bisphosphate metabolic process / phagophore / vacuolar protein processing / positive regulation of vacuole organization / Macroautophagy / cytoplasm to vacuole targeting by the Cvt pathway / nucleophagy / pexophagy / protein localization to phagophore assembly site / piecemeal microautophagy of the nucleus / phagophore assembly site membrane / late endosome to vacuole transport / phosphatidylinositol-3-phosphate binding / fungal-type vacuole membrane / phagophore assembly site / phosphatidylinositol-4-phosphate binding / phosphatidylinositol-3,5-bisphosphate binding / vacuolar membrane / extrinsic component of membrane / autophagosome assembly / ubiquitin binding / cell periphery / macroautophagy / protein transport / endosome membrane / endosome / protein-containing complex / cytosol
Similarity search - Function
: / PROPPIN / WD40 repeats / WD40 repeat / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily
Similarity search - Domain/homology
Autophagy-related protein 18
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
MethodELECTRON MICROSCOPY / subtomogram averaging / cryo EM / Resolution: 26 Å
AuthorsMann, D. / Fromm, S. / Martinez-Sanchez, A. / Gopaldass, N. / Mayer, A. / Sachse, C.
Funding support Germany, 1items
OrganizationGrant numberCountry
Helmholtz Association Germany
CitationJournal: To Be Published
Title: Cryo-EM structures of Atg18 oligomers reveal a tilted structural scaffold for Atg2 at the isolation membrane
Authors: Mann, D. / Fromm, S.A. / Martinez-Sanchez, A. / Gopaldass, N. / Mayer, A. / Sachse, C.
History
DepositionJul 18, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0Nov 1, 2023Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Autophagy-related protein 18
B: Autophagy-related protein 18
C: Autophagy-related protein 18
D: Autophagy-related protein 18
E: Autophagy-related protein 18
F: Autophagy-related protein 18
G: Autophagy-related protein 18
H: Autophagy-related protein 18


Theoretical massNumber of molelcules
Total (without water)440,3668
Polymers440,3668
Non-polymers00
Water0
1


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

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Components

#1: Protein
Autophagy-related protein 18 / Cytoplasm to vacuole targeting protein 18 / Needed for premeiotic replication protein 1 / Swollen ...Cytoplasm to vacuole targeting protein 18 / Needed for premeiotic replication protein 1 / Swollen vacuole phenotype protein 1


Mass: 55045.777 Da / Num. of mol.: 8
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Strain: ATCC 204508 / S288c / Gene: ATG18, AUT10, CVT18, NMR1, SVP1, YFR021W / Production host: Escherichia coli (E. coli) / References: UniProt: P43601

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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: Subtomogram average of membrane-bound Atg18 / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 7.2
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 99 % / Chamber temperature: 291 K

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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: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 0 nm / Nominal defocus min: 0 nm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 3 e/Å2 / Avg electron dose per subtomogram: 120 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k)
EM imaging opticsPhase plate: VOLTA PHASE PLATE

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Processing

EM software
IDNameCategory
7UCSF ChimeraXmodel fitting
9cryoSPARCinitial Euler assignment
10RELIONfinal Euler assignment
11RELIONclassification
12RELION3D reconstruction
13PHENIXmodel refinement
CTF correctionType: NONE
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 26 Å / Resolution method: OTHER / Num. of particles: 8300 / Algorithm: BACK PROJECTION / Details: Mask-less FDR-FSC at 1% FDR / Symmetry type: POINT
EM volume selectionNum. of tomograms: 16 / Num. of volumes extracted: 8300
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Details: Fitted 4x dimer "T" from Atg18 filament structure to interpret the density as a tetramer of dimers

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