NUCLEOPROTEIN STRUCTURES AND GENOME DUPLICATION

理解DNA复制的受控启动机制 这在很大程度上仍然是细胞生物学中一个基本且尚未解决的问题。我们的 长期目标是通过以下方式为解决这一问题做出贡献 对大肠杆菌R6K(PR6K)进行了研究。的主要焦点 我们在当前授权期内的研究将是生物化学 PR6K-激活因子和复制抑制因子的活性分析 编码的启动子蛋白pi和SO-2作用的测定 在复制中称为增强子区域。因为它的伽马或里 PI蛋白结合具有许多结构特征，起源于 许多其他的质粒，拟议的研究可能会有广泛的 生物学意义。然而，重要的是要注意到伽马 ORI在许多方面也不同于其他起源。因此，我们的 研究可能阐明控制DNA复制的新机制。 我们在这项建议中的具体目标是： SA#1。确定以下活动的最小序列要求 体内的Gamma Ori复制。 SA#2.为了在体外表征伽马源的复制特性 带有和不带有增强子序列的模板。 SA#3.以确定复制增强器如何工作。 SA#4.阐明pi蛋白在启动启动中的作用 使用wt和突变的pi蛋白的频率。

Understanding the mechanism of controlled initiation of DNA replication remains a fundamental and largely unsolved problem in cell biology. Our long-term goal is to contribute to the solution of this problem by investigating Escherichia coli plasmid R6K (pR6K). The major focus of our research during the current granting period will be the biochemical analysis of the activator and replication inhibitor activities of pR6K- encoded initiator protein pi and the determination of the role of so- called enhancer region in replication. Because the gamma ori to which pi proteins binds shares a number of structural features with origins of many other plasmids, the proposed studies are likely to have broad biological significance. However, it is important to note that the gamma ori also differs from the other origins in many respects. Thus, our studies may elucidate novel mechanisms of control of DNA replication. Our specific aims in this proposal are: SA#1. To determine minimal sequence requirements for the activity of gamma ori replication in vivo. SA#2. To characterize in vitro the replication properties of gamma ori templates with and without the enhancer sequences. SA#3. To determine how the replication enhancer works. SA#4. To elucidate the role of pi protein in setting initiation frequency by using wt and mutant pi proteins.