Cookies on this website

We use cookies to ensure that we give you the best experience on our website. If you click 'Accept all cookies' we'll assume that you are happy to receive all cookies and you won't see this message again. If you click 'Reject all non-essential cookies' only necessary cookies providing core functionality such as security, network management, and accessibility will be enabled. Click 'Find out more' for information on how to change your cookie settings.

During exocytosis a four-helical coiled coil is formed between the three SNARE proteins syntaxin, synaptobrevin and SNAP-25, bridging vesicle and plasma membrane. We have investigated the assembly pathway of this complex by interfering with the stability of the hydrophobic interaction layers holding the complex together. Mutations in the C-terminal end affected fusion triggering in vivo and led to two-step unfolding of the SNARE complex in vitro, indicating that the C-terminal end can assemble/disassemble independently. Free energy perturbation calculations showed that assembly of the C-terminal end could liberate substantial amounts of energy that may drive fusion. In contrast, similar N-terminal mutations were without effects on exocytosis, and mutations in the middle of the complex selectively interfered with upstream maturation steps (vesicle priming), but not with fusion triggering. We conclude that the SNARE complex forms in the N- to C-terminal direction, and that a partly assembled intermediate corresponds to the primed vesicle state.

Original publication

DOI

10.1038/sj.emboj.7601003

Type

Journal article

Journal

EMBO J

Publication Date

08/03/2006

Volume

25

Pages

955 - 966

Keywords

Amino Acid Sequence, Amino Acid Substitution, Animals, Binding Sites, Cell Membrane, Chromaffin Cells, Circular Dichroism, Electrophysiology, Exocytosis, Membrane Fusion, Mice, Mice, Knockout, Molecular Sequence Data, Mutation, Qa-SNARE Proteins, R-SNARE Proteins, Secretory Vesicles, Sequence Homology, Amino Acid, Synaptosomal-Associated Protein 25