DEEPN as an Approach for Batch Processing of Yeast 2-Hybrid Interactions.
DEEPN as an Approach for Batch Processing of Yeast 2-Hybrid Interactions.
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DEEPN 作为酵母 2-杂交相互作用的批量处理方法。
DOI：
10.1016/j.celrep.2016.08.095
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发表时间：
2016
期刊：
影响因子：
8.8
通讯作者：
Piper,RobertC
中科院分区：
文献类型：
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作者：
Pashkova,Natasha;Peterson,TabithaA;Krishnamani,Venkatramanan;Breheny,Patrick;Stamnes,Mark;Piper,RobertC
我们采用酵母双杂交试验，通过使用称为DEEPN（动态富集蛋白质网络评估）的策略，在单个筛选中同时发现多个瞬时蛋白质相互作用。这种方法结合了高通量DNA测序和计算，以跟踪编码相互作用伴侣的质粒群体之间的竞争。为了证明DEEPN的能力，我们确定了广泛的泛素结合蛋白，包括我们生物化学验证的相互作用。为了证明DEEPN的特异性，我们表明DEEPN允许同时比较多个诱饵蛋白之间的候选相互作用，允许识别差异相互作用。该特征用于识别区分Rab5的GTP结合构象和GDP结合构象的相互作用物。
We adapted the yeast 2-hybrid assay to simultaneously uncover multiple transient protein interactions within a single screen by using a strategy termed DEEPN (dynamic enrichment for evaluation of protein networks). This approach incorporates high-throughput DNA sequencing and computation to follow competition among a plasmid population encoding interacting partners. To demonstrate the capacity of DEEPN, we identify a wide range of ubiquitin-binding proteins, including interactors that we verify biochemically. To demonstrate the specificity of DEEPN, we show that DEEPN allows simultaneous comparison of candidate interactors across multiple bait proteins, allowing differential interactions to be identified. This feature was used to identify interactors that distinguish between GTP- and GDP-bound conformations of Rab5.