Vidal M, Brachmann R K, Fattaey A, Harlow E, Boeke J D
Massachusetts General Hospital Cancer Center, Charlestown 02129, USA.
Proc Natl Acad Sci U S A. 1996 Sep 17;93(19):10315-20. doi: 10.1073/pnas.93.19.10315.
Macromolecular interactions define many biological phenomena. Although genetic methods are available to identify novel protein-protein and DNA-protein interactions, no genetic system has thus far been described to identify molecules or mutations that dissociate known interactions. Herein, we describe genetic systems that detect such events in the yeast Saccharomyces cerevisiae. We have engineered yeast strains in which the interaction of two proteins expressed in the context of the two-hybrid system or the interaction between a DNA-binding protein and its binding site in the context of the one-hybrid system is deleterious to growth. Under these conditions, dissociation of the interaction provides a selective growth advantage, thereby facilitating detection. These methods referred to as the "reverse two-hybrid system" and "reverse one-hybrid system" facilitate the study of the structure-function relationships and regulation of protein-protein and DNA-protein interactions. They should also facilitate the selection of dissociator molecules that could be used as therapeutic agents.
大分子相互作用定义了许多生物学现象。尽管有遗传方法可用于鉴定新的蛋白质 - 蛋白质和DNA - 蛋白质相互作用，但迄今为止尚未描述用于鉴定解离已知相互作用的分子或突变的遗传系统。在此，我们描述了在酿酒酵母中检测此类事件的遗传系统。我们构建了酵母菌株，其中在双杂交系统背景下表达的两种蛋白质之间的相互作用，或在单杂交系统背景下DNA结合蛋白与其结合位点之间的相互作用对生长有害。在这些条件下，相互作用的解离提供了选择性生长优势，从而便于检测。这些方法被称为“反向双杂交系统”和“反向单杂交系统”，有助于研究蛋白质 - 蛋白质和DNA - 蛋白质相互作用的结构 - 功能关系及调控。它们还应有助于选择可作为治疗剂的解离分子。
