Interrogating yeast surface-displayed human proteome to identify small molecule-binding proteins
Interrogating yeast surface-displayed human proteome to identify small molecule-binding proteins
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DOI：
10.1074/mcp.m700223-mcp200
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发表时间：
2007-11-01
影响因子：
7
通讯作者：
Liu, Bin
中科院分区：
文献类型：
--
作者：
Bidlingmaier, Scott;Liu, Bin
识别与小分子相互作用的蛋白质通常是理解细胞信号通路或药物作用分子机制的一个具有挑战性的步骤。在本报告中，我们描述了在酵母细胞表面展示人类蛋白质片段的文库的构建，并证明了这些文库在小分子/蛋白质相互作用研究中的实用性。该文库用于选择对磷脂酰肌醇 4,5-二磷酸 (PtdIns(4,5)P-2) 和磷脂酰肌醇 3,4,5-三磷酸 (PtdIns(3,4,5)P-3) 具有亲和力的蛋白质片段。我们回收了编码 pleckstrin 同源结构域、磷酸酪氨酸结合结构域和载脂蛋白 H 片段的 cDNA 插入片段。 pleckstrin 同源结构域和磷酸酪氨酸结合结构域是已知的磷脂酰肌醇结合结构域，证明了我们方法的有效性。载脂蛋白 H 与 PtdIns(4,5) P-2 和 PtdIns(3,4,5) P-3 的结合以前尚未报道过，因此代表了新的相互作用。我们期望该方法将普遍适用于小分子/蛋白质相互作用的研究，并可能有助于细胞信号通路和药物作用或毒性机制的研究。
Identifying proteins that interact with small molecules is often a challenging step in understanding cellular signaling pathways or molecular mechanisms of drug action. In this report, we describe the construction of libraries displaying human protein fragments on the surface of yeast cells and demonstrate the utility of these libraries for the study of small molecule/protein interactions. The libraries were used to select protein fragments with affinity for the phosphatidylinositides phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P-2) and phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P-3). We recovered cDNA inserts encoding pleckstrin homology domains, a phosphotyrosine-binding domain, and a fragment of apolipoprotein H. The pleckstrin homology and phosphotyrosine-binding domains are known phosphatidylinositide-binding domains, demonstrating the effectiveness of our approach. Binding of apolipoprotein H to PtdIns(4,5) P-2 and PtdIns(3,4,5) P-3 has not been reported previously and thus represents novel interactions. We expect that this method will be generally applicable to the study of small molecule/protein interactions and may facilitate the study of cellular signaling pathways and mechanisms of drug action or toxicity.