eLife
eLife
4 years ago

Uncoupling of dynamin polymerization and GTPase activity revealed by the conformation-specific nanobody Dynab

Sandrine Moutel, Jason Ecard, Valentina Galli, Rafael Sebastian, Franck Perez, Aurélien Roux
Dynamin is a large GTPase that forms a helical collar at the neck of endocytic pits, and catalyzes membrane fission (1, 2). Dynamin fission reaction is strictly dependent on GTP hydrolysis, but how fission is mediated is still debated (3): GTP energy could be spent in membrane constriction required for fission, or in disassembly of the dynamin polymer to trigger fission. To follow dynamin GTP hydrolysis at endocytic pits, we generated a conformation-specific nanobody called dynab, that binds preferentially to the GTP hydrolytic state of dynamin-1. Dynab allowed us to follow the GTPase activity of dynamin-1 in real-time. We show that in fibroblasts, dynamin GTP hydrolysis occurs as stochastic bursts, which are randomly distributed relatively to the peak of dynamin assembly. Thus, dynamin disassembly is not coupled to GTPase activity, supporting that the GTP energy is primarily spent in constriction.

Publisher URL: https://elifesciences.org/articles/25197

DOI: 10.7554/eLife.25197

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