MCM PROTEIN FUNCTION DURING EUKARYOTIC DNA REPLICATION

真核 DNA 复制过程中的 MCM 蛋白功能

• 批准号: 2734872
• 负责人: Stephen P. Bell
• 金额: $ 22.38万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-02-01 至 2003-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2734872
• 关键词: DNA binding protein; DNA replication; Saccharomyces cerevisiae; adenosine triphosphate; cell cycle; fungal genetics; fungal proteins; gene mutation; laboratory mouse; molecular cloning; nucleic acid sequence; protein structure function; site directed mutagenesis; tissue /cell culture

The timely and accurate replication of the genome is essential to the normal proliferation of all eukaryotic cells. Accordingly, the initiation of DNA replication is carefully coordinated with the progress of the cell cycle. The long term objective of this proposal is to determine the events that direct the initiation of replication and how these factors then contribute to the elongation phases of DNA replication. The MCM proteins are a family of six related proteins that play a central role in the replication process in the yeast S. cerevisiae. Recent studies indicate that these proteins are among the earliest factors assembled at origins of replication (prior to the initiation of DNA replication) but also participate in the elongation phase of the replication process. These and other findings suggest that the MCM proteins act as the replicative DNA helicase in eukaryotic cells. This hypothesis is tested in this proposal by addressing the genetic and biochemical properties of the MCM proteins. Specifically Dr. Bell will: Identify conditional mutants in MCM genes that distinguish between the initiation and elongation functions of these proteins. The resulting mutants will be characterized for their effects on the association of the MCM and other proteins with origin and non-origin DNA sequences. Determine the requirement for origin DNA unwinding to assemble MCM proteins at the origin and identify similar elements in other origins of replication. Determine the biochemical properties of purified MCM proteins with particular attention on their ability to bind and hydrolyze ATP and to act as a DNA helicase. Based on previous studies, any new understanding of the fundamental mechanisms of eukaryotic DNA replication in yeast will be readily translated to our understanding of the same process in human cells. The rapid progress that can be made in S. cerevisiae make it a ideal organism to perform these studies. New understanding of these proteins will lead to strong candidate targets for novel anti-fungal compounds. In addition, the understanding of the yeast MCM proteins gained in these studies will direct studies to identify inhibitors of the human analogs of these proteins, which are strong candidate targets for novel chemotherapeutic compounds based on the success of previous DNA replication inhibitors in chemotherapeutic regimens.

基因组的及时和准确复制对于 所有真核细胞的正常增殖。因此， DNA复制的启动与进程密切协调 对细胞周期的影响。这项建议的长期目标是 确定指导复制启动的事件及其方式 这些因素导致了DNA的延伸期。 复制。MCM蛋白是一个由六种相关蛋白组成的家族， 在酵母菌的复制过程中发挥核心作用。 酿酒。最近的研究表明，这些蛋白质是 最早的因素在复制的起始处聚集(在 DNA复制的启动)，但也参与了延伸 复制过程的阶段。这些和其他发现表明， MCM蛋白在真核生物中起复制DNA解旋酶的作用 细胞。这一假设在本提案中得到了验证，方法是解决 MCM蛋白的遗传和生化特性。特指 贝尔博士将： 识别MCM基因中的条件突变，以区分 这些蛋白质的起始和延伸功能。由此产生的 突变体的特征是它们对关联的影响 MCM和其他具有起源和非起源DNA序列的蛋白质。 确定MCM组装的起始DNA解离要求 蛋白质的来源，并确定其他来源的类似元素 复制的结果。 测定纯化的MCM蛋白的生化性质 特别注意它们结合和水解三磷酸腺苷和 发挥DNA解旋酶的作用。 基于之前的研究，任何对根本的新理解 真核DNA在酵母中复制的机制将很容易被 转化为我们对人类细胞中同样过程的理解。这个 在酿酒酵母中可以取得的快速进展使其成为理想的 进行这些研究的生物。对这些蛋白质的新认识 将导致新的抗真菌化合物的强有力的候选目标。 此外，对酵母MCM蛋白的了解在 这些研究将指导研究以确定人类的抑制物 这些蛋白质的类似物，它们是新的强有力的候选靶点 基于前一代DNA成功的化疗化合物 化疗方案中的复制抑制药。