Fluorescent Nucleosides and Oligonucleotides

描述（由申请人提供）：拟议计划的目标是设计，合成和实施新的荧光核苷类似物作为核酸结构，动力学和识别的探针。指导拟议工作的主要标准是保持与天然核碱基的最高可能的结构相似性，将发射转移到更长的波长，并保持足够的发射量子效率。 该项目的具体目标是：（a）设计“理想的”荧光核苷类似物，这些类似物将具有以下特性：（i）与天然核碱基的高度结构相似性，以忠实地模拟它们的大小和形状，以及杂交和识别性质，（ii）红移吸收光谱，以最小化与天然碱基的吸收的重叠，（iii）可接受的发射量子效率和长发射波长（优选在可见光范围内）;（B）设计所需核碱基类似物的简明合成方法;（c）检查新核苷的生物物理特性;（d）将最有希望的寡核苷酸转化为亚磷酰胺，用于自动化DNA和RNA合成;（e）将荧光核碱基掺入寡核苷酸;（f）检查修饰的寡核苷酸的生物物理特性;和（g）开发利用新开发的新型核苷的基于荧光的测定。 对核酸修饰的认识增加（例如，基因表达中的甲基化（甲基化）及其对疾病的影响，以及耐药病原体的迅速出现，使得有必要开发新的工具来有效研究基本过程和发现作为新药的先导化合物。成功的类似物将被应用到新的基于荧光的测定中。这些试验将有助于：（i）研究在发育、遗传疾病和癌症中起关键作用的核酸修饰酶（例如DNA甲基转移酶），（ii）发现由荧光TAR构建体（一种基本的病毒调节元件）辅助的新型抗HIV剂，以及（iii）发现由荧光A位点类似物辅助的靶向细菌核糖体的新型抗生素。这些研究将促进对关键生物过程的基本理解，并将对改善人类健康产生长期影响。

DESCRIPTION (provided by applicant): The goal of the proposed program is to design, synthesize and implement new fluorescent nucleoside analogs as probes for nucleic acids structure, dynamics and recognition. The main criteria directing the proposed work are to maintain the highest possible structural similarity to the natural nucleobases, to shift the emission to longer wavelengths, and to retain an adequate emission quantum efficiency. The specific aims of this project are: (a) To design "ideal" fluorescent nucleoside analogs that will possess the following characteristics: (i) A high structural similarity to the native nucleobases to faithfully mimic their size and shape, as well as hybridization and recognition properties, (ii) A red shifted absorption spectrum to minimize overlap with the absorption of the natural bases, (iii) Respectable emission quantum efficiency and a long emission wavelength (preferably in the visible range); (b) To devise concise synthetic approaches to the desired nucleobase analogs; (c) To examine the photophysical characteristics of the new nucleosides; (d) To convert the most promising ones into phosphoramidites for automated DNA and RNA synthesis; (e) To incorporate the fluorescent nucleobases into oligonucleotides; (f) To examine the photophysical characteristics of the modified oligonucleotides; and (g) To develop fluorescence-based assays that utilize the newly developed novel nucleosides. The increased appreciation for nucleic acids modification (e.g., methylation) in gene expression and its impact on diseases, as well as the rapid emergence of resistant pathogens, necessitate the development of new tools for the effective study of fundamental processes and the discovery of lead compounds as novel drugs. The successful analogs will be implemented into novel fluorescence-based assays. These assays will facilitate: (i) the study of nucleic acids modifying enzymes (e.g. DNA methyl transferases) that play crucial roles in development, genetic diseases and cancers, (ii) the discovery of novel anti-HIV agents assisted by a fluorescent TAR construct, an essential viral regulatory element, and (iii) the discovery of novel antibiotics targeted at the bacterial ribosome assisted by a fluorescent A-site analog. These investigations will advance the fundamental understanding of key biological processes and will have long-term impact on improving human health.