Genetic Dissection of the Specificity of Leucine Zipper Dimerization

该研究应用经典的噬菌体遗传学方法筛选与酵母转录因子Gcn4的亮氨酸拉链相关的大量组合一级结构文库。这些研究的目的是定义可以与野生型GCN4亮氨酸拉链形成异二聚体的序列，并表征异二聚化的主要序列要求。氨基酸侧链在蛋白质二聚过程中的作用将通过随机改变一个简单的亮氨酸拉链结构基序的初级序列来评估。在基于遗传学的系统中，将根据二聚体的能力来筛选每个新结构(将有数百万个这样的结构)的功能意义。两种类型的群体将被测序：(1)那些可以与野生型GCN4亮氨酸拉链形成异二聚体的分子，以及(2)那些形成稳定的同源二聚体但不形成异二聚体的分子(与野生型亮氨酸拉链)。这样的研究将对多肽-多肽、蛋白质-蛋白质相互作用中涉及的结构限制有相当大的洞察力。

The proposed studies apply classical phage genetic approaches to the screening of massive combinatorial primary structure libraries related to the leucine zipper of yeast transcription factor GCN4. The object of these studies is to define sequences which can form heterodimers with wild type GCN4 leucine zippers, and to characterize primary sequence requirements for heterodimerization. %%% The roles of amino acid side chains in protein dimerization processes will be evaluated by altering the primary sequence randomly of a simple leucine zipper structural motif. The functional significance of each new structure (there will be millions of such structures) will be screened, in a genetics based system, in terms of ability to dimerize. Two types of populations will be sequenced: (1) those molecules which can form heterodimers with the wild-type GCN4 leucine zipper, and (2) those molecules which form stable homodimers, but not heterodimers (with the wild- type leucine zipper). Such studies will shed considerable insight into the structural constraints involved in peptide-peptide, protein-protein interactions.