Sequence-Specific Incorporation of Functionalized Bases into RNA

将功能化碱基序列特异性掺入 RNA

• 批准号: 8907307
• 负责人: Christopher Switzer
• 金额: $ 3.1万
• 依托单位: Individual Award
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-08-15 至 1991-02-28
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=8907307&HistoricalAwards=false
• 关键词: Sequence; Specific; Incorporation; Functionalized; Bases

This proposal for research on "Sequence Specific Incorporation of Functionalized Bases into RNA" between Dr. Christopher Y. Switzer and Professor Steven A. Benner of the Eidgenoessische Technische Hochschule, Zurich, Switzerland, is sponsored by NSF. Ribonucleotides are involved in cell division, genetic regulation, viral growth, and tumor transformation. The recent discovery of catalytic RNA and its ability to serve as an RNA polymerase has resurrected early ideas that the first biological catalysts were RNA molecules. There is little information regarding the confor- mation of these molecules in solution. The investigation of RNA tertiary and quaternary structure in such activities as RNA splicing, tRNA maturation, and translation has been limited by the lack of flexibility available with existing RNA synthesis tech- nologies. Unnatural bases containing structural probes (photo- affinity labels, spin-labels), if incorporated at specific sites, would facilitate advances in these areas. A nucleotide base pair distinct from those of A/T(U) and G/C that is incorporated into RNA by a polymerase could allow such sequence-specific introduction of structural probes. The goal of the proposed work is to synthesize the iso-cytidine bases carrying functional groups at the 5-position, and to incorporate them enzymatically into RNA under the direction of a DNA template containing d-iso-G. The iso-C/iso-G base pair has a Watson-Crick hydrogen bonding pattern different from the A/T(U) and G/C base pairs. Because of this different hydrogen bonding pattern, a RNA polymerase should faithfully incorporate the iso-C derivatives opposite only to d-iso-G. These functionalized bases will assist in experiments to probe RNA tertiary and quaternary structure.

瑞士苏黎世Eidgenessische Technische Hochschule的Christopher Y.Switzer博士和Steven A.Benner教授提出的关于“Sequence Special InCorporation of Functionalized Bases to RNA”的研究建议由NSF赞助。核糖核苷酸参与细胞分裂、基因调控、病毒生长和肿瘤转化。最近发现的催化RNA及其作为RNA聚合酶的能力，重新唤醒了早期的想法，即第一个生物催化剂是RNA分子。有关这些分子在溶液中的构象的信息很少。由于现有的RNA合成技术缺乏灵活性，在RNA剪接、tRNA成熟和翻译等活动中对RNA三级和四级结构的研究一直受到限制。含有结构探针(光亲和标记、自旋标记)的非天然碱基，如果结合在特定的位置，将促进这些领域的进展。通过聚合酶结合到RNA中的不同于A/T(U)和G/C的核苷酸碱基对可以允许这种序列特异性的结构探针的引入。这项工作的目标是合成在5位带有官能团的异胞苷碱基，并在含有d-iso-G的DNA模板的指导下，将它们酶法结合到RNA中。Iso-C/iso-G碱基对具有不同于A/T(U)和G/C碱基对的Watson-Crick氢键模式。由于这种不同的氢键模式，RNA聚合酶应该忠实地结合与d-iso-G相反的iso-C衍生物。这些官能化的碱基将有助于探索RNA三级和四级结构的实验。