+Open data
-Basic information
Entry | Database: PDB / ID: 8afy | ||||||
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Title | Subtomogram average of membrane-bound Atg18 oligomers | ||||||
Components | Autophagy-related protein 18 | ||||||
Keywords | MEMBRANE 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 / pexophagy / nucleophagy ...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 / pexophagy / nucleophagy / protein localization to phagophore assembly site / piecemeal microautophagy of the nucleus / phagophore assembly site membrane / late endosome to vacuole transport / phosphatidylinositol-3-phosphate binding / phagophore assembly site / fungal-type vacuole membrane / 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 | ||||||
Biological species | Saccharomyces cerevisiae (brewer's yeast) | ||||||
Method | ELECTRON MICROSCOPY / subtomogram averaging / cryo EM / Resolution: 26 Å | ||||||
Authors | Mann, D. / Fromm, S. / Martinez-Sanchez, A. / Gopaldass, N. / Mayer, A. / Sachse, C. | ||||||
Funding support | Germany, 1items
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Citation | Journal: 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 |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8afy.cif.gz | 610.6 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8afy.ent.gz | 475.3 KB | Display | PDB format |
PDBx/mmJSON format | 8afy.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/af/8afy ftp://data.pdbj.org/pub/pdb/validation_reports/af/8afy | HTTPS FTP |
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-Related structure data
Related structure data | 15412MC 8afqC 8afwC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | 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
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: subtomogram averaging |
-Sample preparation
Component | Name: Subtomogram average of membrane-bound Atg18 / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT |
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Molecular weight | Experimental value: NO |
Source (natural) | Organism: Saccharomyces cerevisiae (brewer's yeast) |
Source (recombinant) | Organism: Escherichia coli (E. coli) |
Buffer solution | pH: 7.2 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Instrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 99 % / Chamber temperature: 291 K |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: TFS KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 0 nm / Nominal defocus min: 0 nm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron 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 optics | Phase plate: VOLTA PHASE PLATE |
-Processing
EM software |
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CTF correction | Type: NONE | |||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | |||||||||||||||||||||
3D reconstruction | Resolution: 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 selection | Num. of tomograms: 16 / Num. of volumes extracted: 8300 | |||||||||||||||||||||
Atomic model building | Protocol: RIGID BODY FIT / Space: REAL Details: Fitted 4x dimer "T" from Atg18 filament structure to interpret the density as a tetramer of dimers |