SITE SPECIFIC ENERGETICS OF PROTEIN/DNA INTERACTIONS

这项研究计划的目标是阐明 由蛋白质相互作用介导的基因控制过程， DNA上的特定位点。该方案的组成部分包括：（a）发展 研究位点特异性蛋白质-DNA相互作用的新技术， (b)特定系统中的功能能量学实验研究，（c） 能量性质与结构特征的相关性 相互作用的分子，以及（d）物理- 基因控制的化学模型 我们计划扩展我们以前对阻遏物-操纵基因调控的研究 系统的噬菌体λ，这是一个大家庭的原型， 合作多位点基因控制系统使用的技术 量热法结合定量DNA酶足迹滴定， 我们开发的一种新的低温凝胶转移技术， 详细研究了温度、pH值和盐的影响， 特定位点能量学（包括协同性）上的相互作用 cI和cro阻遏物与DNA操纵基因位点结合的影响。的 结果将为质子的详细作用提供新的见解 结合，盐结合，以及各种非共价力， 监管互动。这些研究将扩展到一系列 缺乏协同性的突变阻遏物。 我们将研究生物之间的合作相互作用的作用 操纵子结合阻遏物的体外和体内突变研究 阻遏物已经改变了协同性，使用物理化学 基因控制的模型，正在开发下，这一计划， research.

The goal of this research program is to elucidate physical mechanisms of gene control processes mediated by the interactions of proteins at specific sites on DNA. Components of the program include: (a) development of new techniques for the study of site-specific protein-DNA interactions, (b) experimental studies of functional energetics in specific systems, (c) correlation of the energetic properties with structural features of the interacting molecules, and (d) development and testing of physical- chemical models for gene control. We plan to extend our previous studies on the repressor-operator control system of bacteriophage lambda, which is a prototype for a large family of cooperative multi-site gene control systems. Using techniques of calorimetry in combination with quantitative DNase footprint titration and a new cryogenic gel shift technique that we have developed, we will carry out a detailed study of the effects of temperature, pH, and salt interactions on the site-specific energetics (including cooperativity effects) of cI and cro repressor binding to the DNA operator sites. The results will provide new insights into the detailed roles of proton binding, salt binding, and the various non-covalent forces responsible for the regulatory interactions. These studies will be extended to a series of mutant repressors that are deficient in cooperativity. We will study the biological role of cooperative interactions between operator-bound repressors by in vitro and in vivo studies of mutant repressors that have altered cooperativity, using a physical-chemical model of gene control that is being developed under this program of research.