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- PDB-8afw: Tube assembly of Atg18-WT -

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Basic information

Entry
Database: PDB / ID: 8afw
TitleTube assembly of Atg18-WT
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 / positive regulation of vacuole organization / vacuolar protein processing / Macroautophagy / glycophagy / autophagy of peroxisome / cytoplasm to vacuole targeting by the Cvt pathway ...regulation of phosphatidylinositol biosynthetic process / PAS complex / 1-phosphatidyl-1D-myo-inositol 3,5-bisphosphate metabolic process / phagophore / positive regulation of vacuole organization / vacuolar protein processing / Macroautophagy / glycophagy / autophagy of peroxisome / cytoplasm to vacuole targeting by the Cvt pathway / nucleophagy / pexophagy / protein localization to phagophore assembly site / phagophore assembly site membrane / late endosome to vacuole transport / piecemeal microautophagy of the nucleus / 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 / autophagy of mitochondrion / autophagosome assembly / ubiquitin binding / cell periphery / macroautophagy / 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 / helical reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsMann, D. / Fromm, S. / Martinez-Sanchez, A. / Gopaldass, N. / Mayer, A. / Sachse, C.
Funding support Germany, 1items
OrganizationGrant numberCountry
Helmholtz Association Germany
CitationJournal: Nat Commun / Year: 2023
Title: Atg18 oligomer organization in assembled tubes and on lipid membrane scaffolds.
Authors: Daniel Mann / Simon A Fromm / Antonio Martinez-Sanchez / Navin Gopaldass / Ramona Choy / Andreas Mayer / Carsten Sachse /
Abstract: Autophagy-related protein 18 (Atg18) participates in the elongation of early autophagosomal structures in concert with Atg2 and Atg9 complexes. How Atg18 contributes to the structural coordination of ...Autophagy-related protein 18 (Atg18) participates in the elongation of early autophagosomal structures in concert with Atg2 and Atg9 complexes. How Atg18 contributes to the structural coordination of Atg2 and Atg9 at the isolation membrane remains to be understood. Here, we determined the cryo-EM structures of Atg18 organized in helical tubes, Atg18 oligomers in solution as well as on lipid membrane scaffolds. The helical assembly is composed of Atg18 tetramers forming a lozenge cylindrical lattice with remarkable structural similarity to the COPII outer coat. When reconstituted with lipid membranes, using subtomogram averaging we determined tilted Atg18 dimer structures bridging two juxtaposed lipid membranes spaced apart by 80 Å. Moreover, lipid reconstitution experiments further delineate the contributions of Atg18's FRRG motif and the amphipathic helical extension in membrane interaction. The observed structural plasticity of Atg18's oligomeric organization and membrane binding properties provide a molecular framework for the positioning of downstream components of the autophagy machinery.
History
DepositionJul 18, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0Nov 1, 2023Provider: repository / Type: Initial release
Revision 1.1May 15, 2024Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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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


Theoretical massNumber of molelcules
Total (without water)220,6324
Polymers220,6324
Non-polymers00
Water00
1
A: Autophagy-related protein 18
B: Autophagy-related protein 18
C: Autophagy-related protein 18
D: Autophagy-related protein 18
x 20


  • defined by author
  • Evidence: electron microscopy, Please include BioMT matrix as follows to reconstruct filament symmetry cells: REMARK 300 REMARK 300 BIOMOLECULE: 1 REMARK 300 SEE REMARK 350 FOR THE AUTHOR PROVIDED ...Evidence: electron microscopy, Please include BioMT matrix as follows to reconstruct filament symmetry cells: REMARK 300 REMARK 300 BIOMOLECULE: 1 REMARK 300 SEE REMARK 350 FOR THE AUTHOR PROVIDED AND/OR PROGRAM REMARK 300 GENERATED ASSEMBLY INFORMATION FOR THE STRUCTURE IN REMARK 300 THIS ENTRY. THE REMARK MAY ALSO PROVIDE INFORMATION ON REMARK 300 BURIED SURFACE AREA. REMARK 350 REMARK 350 COORDINATES FOR A COMPLETE MULTIMER REPRESENTING THE KNOWN REMARK 350 BIOLOGICALLY SIGNIFICANT OLIGOMERIZATION STATE OF THE REMARK 350 MOLECULE CAN BE GENERATED BY APPLYING BIOMT TRANSFORMATIONS REMARK 350 GIVEN BELOW. BOTH NON-CRYSTALLOGRAPHIC AND REMARK 350 CRYSTALLOGRAPHIC OPERATIONS ARE GIVEN. REMARK 350 REMARK 350 BIOMOLECULE: 1 REMARK 350 SOFTWARE USED: UCSF CHIMERA 1.15_b42258. REMARK 350 APPLY THE FOLLOWING TO CHAINS: A, C, D, E REMARK 350 BIOMT1 1 1.000000 -0.000000 -0.000000 0.00000 REMARK 350 BIOMT2 1 -0.000000 1.000000 -0.000000 0.00000 REMARK 350 BIOMT3 1 -0.000000 -0.000000 1.000000 0.00000 REMARK 350 BIOMT1 2 -0.000000 1.000000 -0.000000 -0.00000 REMARK 350 BIOMT2 2 -1.000000 -0.000000 -0.000000 540.00000 REMARK 350 BIOMT3 2 -0.000000 -0.000000 1.000000 -174.60000 REMARK 350 BIOMT1 3 -0.939693 0.342020 -0.000000 431.37157 REMARK 350 BIOMT2 3 -0.342020 -0.939693 -0.000000 616.06245 REMARK 350 BIOMT3 3 -0.000000 -0.000000 1.000000 -155.20000 REMARK 350 BIOMT1 4 -0.642788 -0.766044 -0.000000 650.38465 REMARK 350 BIOMT2 4 0.766044 -0.642788 -0.000000 236.72065 REMARK 350 BIOMT3 4 -0.000000 -0.000000 1.000000 -135.80000 REMARK 350 BIOMT1 5 0.500000 -0.866025 -0.000000 368.82686 REMARK 350 BIOMT2 5 0.866025 0.500000 -0.000000 -98.82686 REMARK 350 BIOMT3 5 -0.000000 -0.000000 1.000000 -116.40000 REMARK 350 BIOMT1 6 0.984808 0.173648 -0.000000 -42.78310 REMARK 350 BIOMT2 6 -0.173648 0.984808 -0.000000 50.98691 REMARK 350 BIOMT3 6 -0.000000 -0.000000 1.000000 -97.00000 REMARK 350 BIOMT1 7 0.173648 0.984808 -0.000000 -42.78310 REMARK 350 BIOMT2 7 -0.984808 0.173648 -0.000000 489.01309 REMARK 350 BIOMT3 7 -0.000000 -0.000000 1.000000 -77.60000 REMARK 350 BIOMT1 8 -0.866025 0.500000 -0.000000 368.82686 REMARK 350 BIOMT2 8 -0.500000 -0.866025 -0.000000 638.82686 REMARK 350 BIOMT3 8 -0.000000 -0.000000 1.000000 -58.20000 REMARK 350 BIOMT1 9 -0.766044 -0.642788 -0.000000 650.38465 REMARK 350 BIOMT2 9 0.642788 -0.766044 -0.000000 303.27935 REMARK 350 BIOMT3 9 -0.000000 -0.000000 1.000000 -38.80000 REMARK 350 BIOMT1 10 0.342020 -0.939693 -0.000000 431.37157 REMARK 350 BIOMT2 10 0.939693 0.342020 -0.000000 -76.06245 REMARK 350 BIOMT3 10 -0.000000 -0.000000 1.000000 -19.40000 REMARK 350 BIOMT1 11 0.342020 0.939693 -0.000000 -76.06245 REMARK 350 BIOMT2 11 -0.939693 0.342020 -0.000000 431.37157 REMARK 350 BIOMT3 11 -0.000000 -0.000000 1.000000 19.40000 REMARK 350 BIOMT1 12 -0.766044 0.642788 -0.000000 303.27935 REMARK 350 BIOMT2 12 -0.642788 -0.766044 -0.000000 650.38465 REMARK 350 BIOMT3 12 -0.000000 -0.000000 1.000000 38.80000 REMARK 350 BIOMT1 13 -0.866025 -0.500000 -0.000000 638.82686 REMARK 350 BIOMT2 13 0.500000 -0.866025 -0.000000 368.82686 REMARK 350 BIOMT3 13 -0.000000 -0.000000 1.000000 58.20000 REMARK 350 BIOMT1 14 0.173648 -0.984808 -0.000000 489.01309 REMARK 350 BIOMT2 14 0.984808 0.173648 -0.000000 -42.78310 REMARK 350 BIOMT3 14 -0.000000 -0.000000 1.000000 77.60000 REMARK 350 BIOMT1 15 0.984808 -0.173648 -0.000000 50.98691 REMARK 350 BIOMT2 15 0.173648 0.984808 -0.000000 -42.78310 REMARK 350 BIOMT3 15 -0.000000 -0.000000 1.000000 97.00000 REMARK 350 BIOMT1 16 0.500000 0.866025 -0.000000 -98.82686 REMARK 350 BIOMT2 16 -0.866025 0.500000 -0.000000 368.82686 REMARK 350 BIOMT3 16 -0.000000 -0.000000 1.000000 116.40000 REMARK 350 BIOMT1 17 -0.642788 0.766044 -0.000000 236.72065 REMARK 350 BIOMT2 17 -0.766044 -0.642788 -0.000000 650.38465 REMARK 350 BIOMT3 17 -0.000000 -0.000000 1.000000 135.80000 REMARK 350 BIOMT1 18 -0.939693 -0.342020 -0.000000 616.06245 REMARK 350 BIOMT2 18 0.342020 -0.939693 -0.000000 431.37157 REMARK 350 BIOMT3 18 -0.000000 -0.000000 1.000000 155.20000 REMARK 350 BIOMT1 19 -0.000000 -1.000000 -0.000000 540.00000 REMARK 350 BIOMT2 19 1.000000 -0.000000 -0.000000 -0.00000 REMARK 350 BIOMT3 19 -0.000000 -0.000000 1.000000 174.60000 REMARK 350 BIOMT1 20 0.939693 -0.342020 -0.000000 108.62843 REMARK 350 BIOMT2 20 0.342020 0.939693 -0.000000 -76.06245 REMARK 350 BIOMT3 20 -0.000000 -0.000000 1.000000 194.00000
  • 4.41 MDa, 80 polymers
  • Search similar-shape structures of this assembly by Omokage search (details)
Theoretical massNumber of molelcules
Total (without water)4,412,63480
Polymers4,412,63480
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation19

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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: 55157.930 Da / Num. of mol.: 4
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: FILAMENT / 3D reconstruction method: helical reconstruction

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Sample preparation

ComponentName: Filament assembly of Atg18-WT / 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 FIELD / Nominal defocus max: 3000 nm / Nominal defocus min: 1000 nm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 90 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of real images: 1812

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Processing

EM software
IDNameCategory
1cryoSPARCparticle selection
4cryoSPARCCTF correction
7PyMOLmodel fitting
9cryoSPARCinitial Euler assignment
10cryoSPARCfinal Euler assignment
11cryoSPARCclassification
12cryoSPARC3D reconstruction
13PHENIXmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Helical symmertyAngular rotation/subunit: 70 ° / Axial rise/subunit: 19.4 Å / Axial symmetry: C1
3D reconstructionResolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 133981 / Symmetry type: HELICAL
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL / Target criteria: Correlation coefficient
Details: Initial PDB was derived from the Atg18-PR72AA filament structure, subunits were individually docked inside the density and the PR72AA loop was manually modified in Coot

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