Dissociation of synaptotagmin from SNARE complexes

Synaptotagmin是一种含有两个C2结构域（C2A, C2B）的突触囊泡蛋白，是一种被认为与SNARE复合物合作的神经递质释放的Ca2+传感器。然而，使用可溶性片段的实验表明，synaptotagin - snare相互作用需要Ca2+结合到两个C2结构域，Ca2+结合到C2B对于释放更为关键。因此，尚不清楚synaptotagmin如何将Ca2+信号转导到snare介导的胞吐。在这项研究中，我们发现当使用天然蛋白、哺乳动物细胞中表达的蛋白或重组脂质体的细菌表达蛋白时，在37摄氏度下，Ca2+结合到C2B结构域释放syntaxin中的synaptotagmin。结合之前的功能分析，我们的研究结果表明，突触蛋白在传递器释放过程中通过Ca2+与SNARE复合物分离，因此，作为抑制SNARE活性的钳子，直到Ca2+流入。

Synaptotagmin, a synaptic vesicle protein containing two C2 domains (C2A, C2B), is a putative Ca2+ sensor for neurotransmitter release that cooperates with SNARE complexes. Whereas experiments using soluble fragments have shown that synaptotagmin-SNARE interactions require Ca2+ binding to both C2 domains, Ca2+ binding to C2B is more critical for release. Therefore, it is still unknown how synaptotagmin transduces Ca2+ signals to SNARE-mediated exocytosis. In this study, we showed that Ca2+ binding to the C2B domain released synaptotagmin from syntaxin within SNARE complexes at 37 degrees Celsius when using native proteins, proteins expressed in mammalian cells, or bacterial-expressed proteins reconstituted into liposomes. Together with the previous functional analysis, our findings indicate that synaptotagmin dissociates from SNARE complexes by Ca2+ during transmitter release and, thus, acts as a clamp that suppresses SNARE activity until Ca2+ influx.