Biotechnology and Biological Sciences Research Council (BBSRC)
BB/N008391/1
United Kingdom
Citation
Journal: EMBO J / Year: 2021 Title: Multi-modal adaptor-clathrin contacts drive coated vesicle assembly. Authors: Sarah M Smith / Gabrielle Larocque / Katherine M Wood / Kyle L Morris / Alan M Roseman / Richard B Sessions / Stephen J Royle / Corinne J Smith / Abstract: Clathrin-coated pits are formed by the recognition of membrane and cargo by the AP2 complex and the subsequent recruitment of clathrin triskelia. A role for AP2 in coated-pit assembly beyond initial ...Clathrin-coated pits are formed by the recognition of membrane and cargo by the AP2 complex and the subsequent recruitment of clathrin triskelia. A role for AP2 in coated-pit assembly beyond initial clathrin recruitment has not been explored. Clathrin binds the β2 subunit of AP2, and several binding sites have been identified, but our structural knowledge of these interactions is incomplete and their functional importance during endocytosis is unclear. Here, we analysed the cryo-EM structure of clathrin cages assembled in the presence of β2 hinge-appendage (β2HA). We find that the β2-appendage binds in at least two positions in the cage, demonstrating that multi-modal binding is a fundamental property of clathrin-AP2 interactions. In one position, β2-appendage cross-links two adjacent terminal domains from different triskelia. Functional analysis of β2HA-clathrin interactions reveals that endocytosis requires two clathrin interaction sites: a clathrin-box motif on the hinge and the "sandwich site" on the appendage. We propose that β2-appendage binding to more than one triskelion is a key feature of the system and likely explains why assembly is driven by AP2.
History
Deposition
May 20, 2021
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Header (metadata) release
Aug 11, 2021
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Map release
Aug 11, 2021
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Update
Oct 13, 2021
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Current status
Oct 13, 2021
Processing site: PDBe / Status: Released
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Structure visualization
Movie
Surface view with section colored by density value
EMPIAR-10784 (Title: Multi-modal adaptor-clathrin contacts drive coated vesicle assembly Data size: 1.9 Data #1: Hexagonal face subparticles of the 28 mini coat, class 18 [picked particles - single frame - processed])
Entire : Cryo-EM structure of the hexagonal face of the 28 triskelia mini ...
Entire
Name: Cryo-EM structure of the hexagonal face of the 28 triskelia mini clathrin coat complex
Components
Complex: Cryo-EM structure of the hexagonal face of the 28 triskelia mini clathrin coat complex
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Supramolecule #1: Cryo-EM structure of the hexagonal face of the 28 triskelia mini ...
Supramolecule
Name: Cryo-EM structure of the hexagonal face of the 28 triskelia mini clathrin coat complex type: complex / ID: 1 / Parent: 0
Source (natural)
Organism: Sus scrofa (pig)
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Experimental details
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Structure determination
Method
cryo EM
Processing
single particle reconstruction
Aggregation state
particle
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Sample preparation
Buffer
pH: 6.4
Vitrification
Cryogen name: ETHANE-PROPANE / Chamber humidity: 90 % / Chamber temperature: 277.15 K / Instrument: LEICA EM GP Details: 3 uL of sample applied to a grid and blotted for 4.5 s before plunging.
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Electron microscopy
Microscope
FEI TITAN KRIOS
Image recording
Film or detector model: FEI FALCON III (4k x 4k) / Average electron dose: 64.0 e/Å2
Electron beam
Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
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