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目的应用酵母双杂交系统筛选小鼠、大鼠、人类大脑文库中与DRP2相互作用的蛋白,分析中枢神经系统中DRP2分子复合体的组成成分。方法设计3个含有DRP2不同蛋白结构域的诱饵质粒,在完成诱饵质粒的自激活性的鉴定后,对小鼠、大鼠、人类脑MATCHMAKER cDNA文库进行筛选。结果运用DRP2蛋白N-Bait筛选大鼠脑MATCHMAKER cDNA文库,笔者获得了一个阳性文库质粒。该质粒的插入子编码一种SNAP25相互作用的蛋白质Scoilin。Scoilin又被命名为Zwint-1蛋白。同时笔者还确定了这两种相互作用蛋白质的精确结构域:DRP2蛋白以其含有两个Spectrin重复和一个WW蛋白结构域的氨基端与Scoilin(Zwint-1)全蛋白相互作用。结论应用酵母双杂交,笔者于大鼠大脑中找到了DRP2的相互作用的蛋白质Scoilin(Zwint-1),并且确定了这两种相互作用蛋白质的精确结构域。
Objective To screen the proteins interacting with DRP2 in mouse, rat and human brain libraries by yeast two-hybrid system and analyze the components of DRP2 complex in the central nervous system. Methods Three bait plasmids containing different domains of DRP2 were designed. After identifying the self-activation of the bait plasmid, mouse, rat and human brain MATCHMAKER cDNA libraries were screened. Results Using the DRP2 protein N-Bait to screen rat brain MATCHMAKER cDNA library, the author obtained a positive library plasmid. The plasmid insert encodes a SNAP25-interacting protein, Scoilin. Scoilin is also named Zwint-1 protein. The authors also identified the precise domains of these two interacting proteins: the DRP2 protein interacts with Scoilin (Zwint-1) whole protein at its amino terminus containing two Spectrin repeats and one WW protein domain. Conclusion Using yeast two-hybrid assay, we found a protein Scoilin (Zwint-1) that interacts with DRP2 in the rat brain and identified the precise domains of these two interacting proteins.