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）： 该提案旨在利用一系列非天然核苷来探究核酸的结构和功能。受到 Nelson Leonard 的苯扩展腺苷类似物的启发，我们将通过在天然嘌呤和嘧啶支架中引入多样性来改变核碱基的宽度，以及随后的 DNA 螺旋。将吡咯、噻吩或呋喃间隔环掺入杂芳族结构中将提供以前研究的碱基对不可能实现的独特结构优势。这项研究将通过对 DNA 螺旋稳定性的物理方面以及静电和碱基堆积所发挥的作用提供新的见解来解决一些基本的科学问题。它还将提供有关核碱基形状、碱基堆积和静电对 DNA 聚合酶复制的影响的有用数据。我们预测，由于静电表面积增加，以及色散、范德华力和极化率增加，杂芳族间隔环提供的优势将显着有助于增强碱基堆叠。此外，杂芳族间隔环具有与阳离子或与螺旋水合脊柱中存在的水形成氢键的能力。因此，我们还预测 DNA 螺旋稳定性将显着增加。此外，这些延伸碱基具有荧光性，可用于许多应用，例如螺旋形成的自我报告、追踪单点突变以及合成 DNA 微阵列。本研究的初步目的是（i）设计、合成和表征一系列扩展嘧啶和扩展嘌呤核苷； (ii) 将非天然核苷掺入不同长度和组成的 DNA 双链体中； (iii) 通过计算、光谱、热力学和结构表征所产生的螺旋； (iv) 评估它们对 DNA 聚合酶保真度的影响。