权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Mechanism of Triplex DNA Formation ; And Its Application to Antisense

三链DNA形成机制；

基本信息

批准号：
09558090
负责人：
SARAI Akinori
金额：
$ 7.42万
依托单位：
RIKEN Insutitute
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 2000
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-09558090/
关键词：
Triplex DNA Thermodynamics Computer simulation Antisense

项目摘要

In order to understand the mechanism of stability and specificity in triplex DNA formation, we performed experimental and computational analyses. We synthesized DNA with chemical modifications at different locations, and measured thermodynamic quantities such as dissociation constant and kinetic quantities such as association rate constant for the triplex formation. These results showed that the chemical modification, particularly at backbone, affected the thermodynamic and kinetic quantities of triplex formation. We attempted to investigate the molecular mechanism of that effect by computational analysis. We performed quantum-chemical calculations for the modified DNA to obtain partial atomic charges and conformation of DNA. We found that the phosphorothioation of DNA has significant effect on the charges and conformation. We also performed computer simulation of DNA to calculate its average conformation and fluctuation, by using a new algorithm for conformational sampling. The chemical modification affected the average conformation and fluctuation of DNA depending on the location of the modification. These results suggest that the chemical modification may affect the stability and specificity of triplex DNA formation through the changes in structure and property of DNA. In order to understand the biological role of triplex DNA in cell, we made biochemical and functional analyses of MAZ protein, which binds to triplex DNA. This protein acts as a transcription factor and regulates the expression of c-myc gene. We also examined the effect of chemical modification of DNA on the binding and transcriptional activities of an oncogene product, Myb. We found that the modification destabilized the DNA and abolished the Myb binding. The present experimental and computational studies of triplex DNA have laid a basis for the design of antisense DNA, which inhibits expression of genes.

为了了解三链体 DNA 形成的稳定性和特异性机制，我们进行了实验和计算分析。我们合成了在不同位置进行化学修饰的 DNA，并测量了解离常数等热力学量和三链体形成的缔合速率常数等动力学量。这些结果表明化学修饰，特别是主链上的化学修饰，影响三链体形成的热力学和动力学量。我们试图通过计算分析来研究该效应的分子机制。我们对修饰后的 DNA 进行了量子化学计算，以获得 DNA 的部分原子电荷和构象。我们发现DNA的硫代磷酸化对电荷和构象有显着影响。我们还使用一种新的构象采样算法对 DNA 进行了计算机模拟，以计算其平均构象和波动。化学修饰会影响 DNA 的平均构象和波动，具体取决于修饰的位置。这些结果表明化学修饰可能通过DNA结构和性质的变化影响三链体DNA形成的稳定性和特异性。为了了解三链体DNA在细胞中的生物学作用，我们对与三链体DNA结合的MAZ蛋白进行了生化和功能分析。该蛋白作为转录因子并调节 c-myc 基因的表达。我们还研究了 DNA 化学修饰对癌基因产物 Myb 的结合和转录活性的影响。我们发现这种修饰破坏了 DNA 的稳定性并消除了 Myb 结合。目前三链体DNA的实验和计算研究为抑制基因表达的反义DNA的设计奠定了基础。