Yeast Protein-Protein Interaction Assays and Screens
Yeast Protein-Protein Interaction Assays and Screens
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DOI：
10.1007/978-1-61779-154-3_8
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发表时间：
2011-01-01
期刊：
影响因子：
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通讯作者：
Immink, Richard G. H.
中科院分区：
文献类型：
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作者：
de Folter, Stefan;Immink, Richard G. H.
大多数转录因子在蛋白质复合体中发挥作用。因此，关于它们相互作用能力的信息揭示了蛋白质的功能和这种活动背后的分子机制。酵母双杂交GAL4 （Y2H）测定是一种强大的方法来解开和鉴定蛋白质复合物的组成。这种基于体内的系统利用了GAL4转录因子的两个功能蛋白结构域，每个都融合到感兴趣的蛋白质上。当研究中的两种蛋白质相互作用时，转录激活子被重组，报告基因被激活，允许酵母在选择性培养基上生长。本章给出了Y2H文库筛选、定向Y2H筛选、Y2H基质筛选和涉及两种以上蛋白的YnH筛选的方案。
Most transcription factors fulfill their role in protein complexes. As a consequence, information about their interaction capacity sheds light on a protein's function and the molecular mechanism underlying this activity. The yeast two-hybrid GAL4 (Y2H) assay is a powerful method to unravel and identify the composition of protein complexes. This in vivo based system makes use of two functional protein domains of the GAL4 transcription factor, each fused to a protein of interest. Upon interaction between the two proteins under study, a transcriptional activator gets reconstituted and reporter genes get activated, allowing the yeast to grow on selective medium. In this chapter protocols are given for Y2H library screening, directed Y2H screening, Y2H matrix screening, and YnH screening involving more than two proteins.