Unnatural Base Pairs as DNA Bioprobes

作为 DNA 生物探针的非天然碱基对

• 批准号: 6898670
• 负责人: Katherine L Seley-Radtke
• 金额: $ 22.71万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE COUNTY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-05-01 至 2009-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6898670
• 关键词: Unnatural; Base; Pairs; DNA; Bioprobes

DESCRIPTION (provided by applicant): This proposal is aimed at using a series of unnatural nucleosides to interrogate nucleic acid structure and function. Drawing inspiration from Nelson Leonard's benzene-expanded adenosine analogues, we will vary the width of the nucleobases, and subsequently the DNA helix, by introduction of diversity into the natural purine and pyrimidine scaffolds. Incorporation of either a pyrrole, thiophene or furan spacer ring into the heteroaromatic architecture will provide unique structural advantages not possible with previously studied base pairs. This research will address some fundamental scientific questions by providing new insights into the physical aspects of DNA helix stability and the role that electrostatics and base stacking play. It will also provide useful data regarding the effect nucleobase shape, base stacking and electrostatics have on DNA polymerase replication. We predict that the advantages provided by the heteroaromatic spacer rings will contribute significantly to enhanced base stacking, due to increased electrostatic surface area, as well as increased dispersion and van der Waals forces and polarizability. Furthermore, the heteroaromatic spacer rings possess the ability to hydrogen bond with cations or with water present in the spine of hydration of the helix. As a result, we also predict a significant increase in DNA helix stability will be observed. In addition, these extended bases are fluorescent and could prove useful in many applications, such self-reporters of helix formation, tracking single point mutations, as well as in synthetic DNA microarrays. The initial aims of this study are to (i) design, synthesize and characterize a series of extended pyrimidines and expanded purine nucleosides; (ii) to incorporate the unnatural nucleosides into DNA duplexes of varying length and composition; (iii) to computationally, spectroscopically, thermodynamically and structurally characterize the resulting helices; and (iv) evaluate their effect on DNA polymerase fidelity.

描述（由申请人提供）： 该建议旨在使用一系列不自然的核苷来询问核酸的结构和功能。我们将通过将多样性引入天然嘌呤和嘧啶支架中，从纳尔逊·伦纳德（Nelson Leonard）的苯膨胀腺苷类似物中汲取灵感，然后改变核碱基的宽度，然后改变DNA螺旋的宽度。将吡咯，噻吩或呋喃垫圈环掺入异芳族建筑中，将在先前研究的碱基对中不可能提供独特的结构优势。这项研究将通过提供有关DNA螺旋稳定性的物理方面的新见解以及静电和基本堆叠发挥的作用来解决一些基本科学问题。它还将提供有关核酶形状，碱基堆叠和静电对DNA聚合酶复制的有用数据。我们预测，由于静电表面积的增加以及分散体和范德华的力和极化性，异质隔离环提供的优势将对增强的基础堆叠产生显着贡献。此外，杂芳族间隔环具有与阳离子或螺旋脊柱中存在的水氢键的能力。结果，我们还预测将观察到DNA螺旋稳定性的显着增加。此外，这些扩展的碱基是荧光的，并且在许多应用中可能有用，例如螺旋形成的自我重新堆，跟踪单点突变以及合成DNA微阵列。这项研究的最初目的是（i）设计，合成和表征一系列扩展的嘧啶和扩展的嘌呤核苷； （ii）将非天然核苷掺入不同长度和组成的DNA双链体中； （iii）从计算，光谱，热力学和结构上表征所得的螺旋； （iv）评估它们对DNA聚合酶保真度的影响。