National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)
GM108753
United States
Citation
Journal: Sci Adv / Year: 2020 Title: The structures of natively assembled clathrin-coated vesicles. Authors: Mohammadreza Paraan / Joshua Mendez / Savanna Sharum / Danielle Kurtin / Huan He / Scott M Stagg / Abstract: Clathrin-coated vesicles mediate trafficking of proteins and nutrients in the cell and between organelles. Proteins included in the clathrin-coated vesicles (CCVs) category include clathrin heavy ...Clathrin-coated vesicles mediate trafficking of proteins and nutrients in the cell and between organelles. Proteins included in the clathrin-coated vesicles (CCVs) category include clathrin heavy chain (CHC), clathrin light chain (CLC), and a variety of adaptor protein complexes. Much is known about the structures of the individual CCV components, but data are lacking about the structures of the fully assembled complexes together with membrane and in complex with cargo. Here, we determined the structures of natively assembled CCVs in a variety of geometries. We show that the adaptor β2 appendages crosslink adjacent CHC β-propellers and that the appendage densities are enriched in CCV hexagonal faces. We resolve how adaptor protein 2 and other associated factors in hexagonal faces form an assembly hub with an extensive web of interactions between neighboring β-propellers and propose a structural model that explains how adaptor binding can direct the formation of pentagonal and hexagonal faces.
History
Deposition
Mar 30, 2020
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Header (metadata) release
Apr 8, 2020
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Map release
Aug 19, 2020
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Update
Aug 19, 2020
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Current status
Aug 19, 2020
Processing site: RCSB / Status: Released
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Structure visualization
Movie
Surface view with section colored by density value
Model: C-flat-2/2 4C / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 40.0 nm
Vitrification
Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 281 K / Instrument: FEI VITROBOT MARK IV
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Electron microscopy
Microscope
FEI TITAN KRIOS
Image recording
Film or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 800 / Average electron dose: 50.0 e/Å2
Electron beam
Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Type of model: OTHER Details: Ab initio model from Relion on a small subset of particles. The ab initio model was used in a Relion classification on all particles minus mini-coat particles.
Final reconstruction
Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 15.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cisTEM (ver. 1) / Number images used: 6300
Initial angle assignment
Type: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3)
Software - Name: cisTEM (ver. 1) Details: Multiple rounds of 3D classification in cisTEM produced different geometries of clathrin coated vesicles. Each of which was classified until stable subsets were achieved.
FSC plot (resolution estimation)
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