NUCLEOPROTEIN STRUCTURES AND GENOME DUPLICATION

理解DNA复制的受控起始机制 仍然是细胞生物学中一个基本的和大部分未解决的问题。 我们 长期目标是通过以下方式为解决这一问题作出贡献： 研究大肠杆菌质粒R6K（pR6K）。 的主要重点 在目前的资助期内，我们的研究将是生物化学 分析pR6K的激活剂和复制抑制剂活性， 编码的起始蛋白PI和确定的作用， 在复制中称为增强子区域。 因为伽马射线 pi蛋白结合的许多结构特征与 许多其他质粒，拟议的研究可能有广泛的 生物学意义 然而，重要的是要注意，伽马 ori在许多方面也不同于其他起源。 所以我们 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。 为了确定活动的最低顺序要求， 体内γ ori复制。 SA #2。 为了在体外表征γ ori的复制特性， 具有和不具有增强子序列的模板。 SA #3。 以确定复制增强器是如何工作的。 SA #4. 阐明pi蛋白在凝固起始中的作用 频率通过使用wt和突变体pi蛋白。

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.