Structural Basis for the Clamping and Ca2+Activation of SNARE-mediated Fusion by Synaptotagmin

Synapotagmin-1 (Syt1) interacts with both SNARE proteins and lipid membranes to synchronize neurotransmitter release to Ca 2+ -influx. To understand the underlying molecular mechanism, we determined the structure of the Syt1-SNARE complex on lipid membranes using cryo-electron microscopy. Under resting conditions, the Syt1 C2 domains adopt a novel membrane orientation with a Mg 2+ -mediated partial insertion of the aliphatic loops, alongside weak interactions with the anionic lipid headgroups. The C2B domain concurrently binds the SNARE bundle via the ‘primary’ interface and is positioned between the SNAREpins and the membrane. In this configuration, Syt1 is projected to sterically delay the complete assembly of the associated SNAREpins and thus, contribute to clamping fusion. This Syt1-SNARE organization is disrupted upon Ca 2+ -influx as Syt1 reorients into the membrane, allowing the attached SNAREpins to complete zippering and drive fusion. Overall, we find cation (Mg 2+ /Ca 2+ ) dependent membrane interaction is a key determinant of the dual clamp/activator function of Syt1.