Protein-protein interactions are one of the most basic and critical processes underlying biological functions. Thus, identification of the interacting proteins of a protein of interest and further elucidation of the roles of the interactions is critical for understanding the related biological processes. The yeast two-hybrid (Y2H) method is a popular approach for identifying protein-protein interactions. Once interacting proteins are identified, a comparison of the phenotypes of mutants lacking the specific protein-protein interaction with those of the wild-type strain is a powerful tool for uncovering the former interactions' biological significance. However, isolation of such interaction-defective mutants is often laborious. Here, I describe a novel and efficient approach for isolating such mutants that uses the Y2H technique with a modified Y2H vector, and provide an example of how this approach can be used to screen interaction-null/impaired mutants. Because the strategy is simple and the modification of a pre-existing Y2H vector is sufficient for the screening purpose, the same strategy can be applied to any existing two-hybrid system.

中文翻译：

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使用酵母双杂交技术分离无效相互作用/受损的突变体的有效方法。

蛋白质-蛋白质相互作用是生物学功能中最基本和最关键的过程之一。因此，鉴定目的蛋白的相互作用蛋白以及进一步阐明相互作用的作用对于理解相关的生物学过程至关重要。酵母双杂交（Y2H）方法是一种用于识别蛋白质-蛋白质相互作用的流行方法。一旦鉴定出相互作用的蛋白质，将缺乏特异性蛋白质-蛋白质相互作用的突变体的表型与野生型菌株的突变体进行比较，便是揭示先前相互作用的生物学意义的有力工具。然而，分离这种相互作用缺陷的突变体通常是费力的。这里，我描述了一种使用这种Y2H技术和经过修饰的Y2H载体来分离此类突变体的新颖有效的方法，并提供了该方法如何可用于筛选无相互作用/受损的突变体的示例。因为该策略很简单，并且已经存在的Y2H向量的修饰足以满足筛选目的，所以相同的策略可以应用于任何现有的双杂交系统。