Synaptic soluble N-ethylmaleimide-sensitive factor attachment
protein receptors (SNAREs) couple their stepwise folding to fusion
of synaptic vesicles with plasma membranes. In this process, three
SNAREs assemble into a stable four-helix bundle. Arguably, the
first and rate-limiting step of SNARE assembly is the formation of
an activated binary t-SNARE complex on the plasma membrane,
which then zippers with the v-SNARE on the vesicle to drive
membrane fusion. However, the t-SNARE complex readily misfolds
and its structure, stability, and dynamics are elusive. Using
single-molecule force spectroscopy, we modeled synaptic t-SNARE
complex as a parallel three-helix bundle with a small frayed Cterminus.
The helical bundle sequentially folded in an N-terminal
domain (NTD) and a C-terminal domain (CTD) separated by a
central ionic layer, with total unfolding energy of ∼17 kBT. Peptide
binding to the CTD activated the t-SNARE complex to initiate
NTD zippering with the v-SNARE, a mechanism likely shared by
Munc18-1. The NTD zippering then dramatically stabilized the CTD,
facilitating further SNARE zippering. The subtle bidirectional tSNARE
conformational switch was mediated by the ionic layer.
Thus, the t-SNARE complex acts as a switch to enable fast and
controlled SNARE zippering required for synaptic vesicle fusion
and neurotransmission
