G-quadruples Topology-selective Probes

G-四联体拓扑选择性探针

• 批准号: 1518006
• 负责人: Edwin Lewis
• 金额: $ 62.5万
• 依托单位: Mississippi State University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-01 至 2019-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1518006&HistoricalAwards=false
• 关键词: quadruples; Topology; selective; Probes

With this award, the Chemistry of Life Processes Program in the Division of Chemistry is funding Drs. Edwin Lewis from Mississippi State University and Herman Sintim from the University of Maryland to develop new chemical probes for the study of G-quadruplex biology in complex biological environments. G-quadruplexes, uniquely folded non-helical structures formed by short guanine rich sequences in DNA or RNA, have been implicated in the regulation of a number of important life processes, from gene expression and mRNA splicing, to chromosome maintenance, cell proliferation, and viral life cycles. The development of chemical probes having very high affinity for G-quadruplex DNA over double stranded helical DNA, and having the ability to select for specific G-quadruplex folding topologies represents a new direction in biochemistry and molecular biology. The design of the new chemical probes, their synthesis and the biophysical characterization of the probe/G-quadruplex complexes in vitro and in vivo will provide opportunities for graduate students and advanced undergraduates to acquire a diverse set of specialized skills in new compound synthesis and in the use of spectroscopic, microcalorimetric, NMR, mass spectrometry, and molecular biology methods. This project is designed to address the central issue in the in vivo study of G-quadruplex biology; the lack of selective probes that can discriminate between: a) abundant duplex nucleic acids and much less abundant G-quadruplexes; b) DNA G-quadruplexes and RNA G-quadruplexes; c) and among several G-quadruplex topologies or conformers. In addition to these unresolved selectivity issues, most G-quadruplex ligands that have been developed to date have only moderate binding affinities with only a handful of G-quadruplex ligands exhibiting nanomolar Kd values. The principal investigators, recently made a surprising discovery that diminazene (DMZ), a drug that is used in veterinary medicine and believed to act via binding to the minor groove of AT-rich duplexes with a Kd of about 1 micromolar, binds to selected G-quadruplexes with Kds of about 1 nanomolar - the strongest affinity reported so far for a G-quadruplex chemical probe. Based on this discovery, this project will undertake the development of new G-quadruplex chemical probes based on the DMZ scaffold. Various biophysical tools will be employed to study the selectivity of the new chemical probes for different G-quadruplex topologies (e.g. parallel vs. antiparallel, and small loop vs. large loop G-quadruplexes) and types (DNA vs. RNA). The interactions of nucleic acid binding proteins with various probe-stabilized G-quadruplex structures will also be investigated in this project. Information developed in this study is expected to provide tools to study the role of G-quadruplex nucleic acid structures in a number of cellular processes.

有了这个奖项，化学部的生命过程化学计划正在资助来自密西西比州立大学的埃德温刘易斯博士和来自马里兰州大学的赫尔曼辛蒂姆博士开发新的化学探针，用于在复杂的生物环境中研究G-四链体生物学。 G-四链体是由DNA或RNA中富含鸟嘌呤的短序列形成的独特折叠的非螺旋结构，它参与了许多重要的生命过程的调节，从基因表达和mRNA剪接到染色体维持、细胞增殖和病毒生命周期。 与双链螺旋DNA相比，开发对G-四链体DNA具有非常高亲和力的化学探针，并具有选择特定G-四链体折叠拓扑结构的能力，代表了生物化学和分子生物学的新方向。 新的化学探针的设计，它们的合成和生物物理特性的探针/G-四链体复合物在体外和体内将提供机会，研究生和高级本科生获得一套不同的专业技能，在新的化合物合成和使用光谱，微量热，核磁共振，质谱和分子生物学方法。 该项目旨在解决G-四链体生物学体内研究中的核心问题;缺乏可以区分以下各项的选择性探针：a）丰富的双链体核酸和丰度低得多的G-四链体; B）DNA G-四链体和RNA G-四链体; c）以及几种G-四链体拓扑结构或构象异构体。 除了这些未解决的选择性问题之外，迄今为止开发的大多数G-四链体配体仅具有中等的结合亲和力，只有少数G-四链体配体表现出纳摩尔Kd值。 主要研究人员最近做出了一个令人惊讶的发现，即二甲氮烯（DMZ），一种用于兽医学的药物，被认为通过结合富含AT的双链体的小沟发挥作用，Kd约为1微摩尔，结合选定的G-四链体，Kd约为1纳摩尔-迄今为止报道的G-四链体化学探针的最强亲和力。 基于这一发现，本项目将承担基于DMZ支架的新型G-四链体化学探针的开发。 将采用各种生物物理工具来研究新化学探针对不同G-四链体拓扑结构（例如平行与反平行，小环与大环G-四链体）和类型（DNA与RNA）的选择性。核酸结合蛋白与各种探针稳定的G-四链体结构的相互作用也将在本项目中进行研究。 在这项研究中开发的信息，预计将提供工具，研究G-四链体核酸结构在一些细胞过程中的作用。