最近的研究表明PIP2 分子通过与Syntaxin相互作用在膜上形成PIP2微结构区。该微结构区的形成对神经递质的释放十分重要。在本研究计划中，我们将利用X-射线，二维NMR，超分辨显微成像，在体离体膜重组方法等实验技术来研究Syntaxin，Munc18 和PIP2磷脂分子对PIP2 微结构区的调控关系。目前，我们已经发现Munc18和PIP2 存在相互作用，我们将考察PIP2 对Munc18蛋白的空间构象影响。而且，我们将进行Munc18 和PIP2的共结晶工作，试图解析Munc18-PIP2 复合物的分子结构。最后，我们将通过人工膜重组技术在膜上考察PIP2微结构区的形成以及与Munc18，Syntaxin和PIP2磷脂分子的关系。我们希望通过该课题研究提出PIP2微结构区形成的分子模型。

It has been recently demonstrated that the membrane lipid PIP2 interacts with Snare protein Syntaxin to induce membrane sequesting and microdomain formation. The formation of PIP2 microdomains has many advantages. First, accumulation of PIP2 around Syntaxin clusters may function as a docking site and recruit related membrane proteins to the active zone to facilitate assembly of the complete fusion machinery. Second, PIP2 microdomains modulate the membrane environment and low down the energetic requirement for fusion. Third, both Syntaxin and PIP2 microdomains regulate the activities of fusion machinery and increase the membrane fusion efficiency. The mechanism of PIP2 microdomains formation is still unclear. In this project, we will use combined techniques, including X-ray, 2D-NMR, super-resolution microscopy and lipid reconstitution assay to study how PIP2 microdomains formation is regulated by Syntaxin, Munc18 and PIP2 molecules. We will investigate the conformation change of Munc18 by adding PIP2 since we found Munc18 binds PIP2 directly. Furthermore, we will co-crystallize Munc18-PIP2 complex and solve its structure. Last but not least, we will do reconstitution assay to monitor PIP2 microdomains formation on liposome. All of these data will help us raise a new model describing how these factors work together and facilitate Syntaxin clusters and PIP2 microdomains formation.