Helical and Superhelical Structure of DNA

DNA 的螺旋和超螺旋结构

• 批准号: 8807067
• 负责人: James Wang
• 金额: $ 50.6万
• 依托单位: Harvard University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-06-15 至 1993-11-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=8807067&HistoricalAwards=false
• 关键词: Helical; Superhelical; Structure; DNA

The experiments proposed in this renewal application revolve around three primary goals. The first is to design an enzyme for making positively supercoiled DNA. It is based on the thesis that during transcription a positive superhelical debt is built ahead of the replication fork while a negative superhelical tension accumulates behind the replication fork. The situation is dramatic when both the DNA and polymerase are constrained during the act of transcription. The model for such an enzyme is a fusion protein that generates both conditions, that is, a protein containing a sequence-specific DNA binding domain joined to RNA polymerase. The second goal is to analyze chromatin structure in vivo. DNA nucleases will be cloned under the control of inducible regulatory sequences in yeast and in higher eukaryotic cells. To obviate problems in making nucleases in vivo, the strategy is to construct cold sensitive mutants that are inactive at low temperature, but able to cleave DNA in vivo when the medium is shifted to a higher temperature. The third goal is to make supercoiled RNA. The results of the research should enhance our understanding of the structure of genes and how they function at the molecular level in living systems.

在这次更新申请中提出的实验 围绕三个主要目标。 首先是设计一个 制造正超螺旋DNA的酶。 它是基于 在转录过程中，一个正超螺旋 债务是建立在复制叉之前， 超螺旋张力在复制叉后面积聚。 当DNA和聚合酶都被 在转录过程中受到限制。 模型 这样的酶是一种融合蛋白， 条件下，即，含有序列特异性的蛋白质 DNA结合域连接到RNA聚合酶。 第二个目标 是分析活体染色质结构。 DNA核酸酶将 在可诱导调控序列的控制下克隆 在酵母和高等真核细胞中。 解决问题 在体内制造核酸酶时，策略是构建冷的 敏感突变体在低温下不活跃，但 当介质转移到一个合适的温度时， 更高的温度。 第三个目标是制造超螺旋RNA。 研究的结果应该会提高我们的 了解基因的结构及其功能 在生命系统的分子水平上。