Flexible open conformation of the AP-3 complex explains its role in cargo recruitment at the Golgi
| Autor: | Yaping Han, Jannis Schoppe, Heike Stephanowitz, Angela Perz, Amir Apelbaum, Oliver Hofnagel, Jacob Piehler, Fan Liu, Christian Ungermann, Stefan Raunser, Evelyn Schubert, Oliver Birkholz, Erdal Yavavli |
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| Rok vydání: | 2021 |
| Předmět: |
Saccharomyces cerevisiae Proteins
Phosphatidylinositol 4-phosphate Endosome AP adaptor protein Biological Transport Active Golgi Apparatus Saccharomyces cerevisiae Biochemistry symbols.namesake chemistry.chemical_compound Editors' Pick Highlight clathrin Golgi Small GTPase Molecular Biology ARF ADP-ribosylation factor Vesicle Signal transducing adaptor protein ARF Cell Biology Golgi apparatus AP-1 AP-2 AP-3 phosphoinositide Membrane chemistry Multiprotein Complexes symbols Biophysics cryo-EM Guanine nucleotide exchange factor |
| Zdroj: | The Journal of Biological Chemistry |
| ISSN: | 1083-351X |
| Popis: | Vesicle formation at endomembranes requires the selective concentration of cargo by coat proteins. Conserved adapter protein complexes at the Golgi (AP-3), the endosome (AP-1), or the plasma membrane (AP-2) with their conserved core domain and flexible ear domains mediate this function. These complexes also rely on the small GTPase Arf1 and/or specific phosphoinositides for membrane binding. The structural details that influence these processes, however, are still poorly understood. Here we present cryo-EM structures of the full-length stable 300 kDa yeast AP-3 complex. The structures reveal that AP-3 adopts an open conformation in solution, comparable to the membrane-bound conformations of AP-1 or AP-2. This open conformation appears to be far more flexible than AP-1 or AP-2, resulting in compact, intermediate, and stretched subconformations. Mass spectrometrical analysis of the cross-linked AP-3 complex further indicates that the ear domains are flexibly attached to the surface of the complex. Using biochemical reconstitution assays, we also show that efficient AP-3 recruitment to the membrane depends primarily on cargo binding. Once bound to cargo, AP-3 clustered and immobilized cargo molecules, as revealed by single-molecule imaging on polymer-supported membranes. We conclude that its flexible open state may enable AP-3 to bind and collect cargo at the Golgi and could thus allow coordinated vesicle formation at the trans-Golgi upon Arf1 activation. |
| Databáze: | OpenAIRE |
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