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Molecular Population Dynamics of DNA Tetraplexes using Magneto-Optical Tweezers

机译：使用磁光镊子的DNA四联体的分子种群动力学。

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摘要

The most common B-form Deoxyribonucleic acid (DNA) consists of two polymeric nucleic acid chains held together by Watson-Crick base pair (bp) with a right-handed twist at every 10.5 bp. In the nucleus, the DNA is subjected to spatial constraint and hence undergoes supercoiling (sigma) to relieve stress through either overwinding (positive superhelicity), unwinding (negative superhelicity), DNA bending or melting. Similar topological changes observed during many cellular processes such as transcription, replication, etc. have a significant effect on the rate of these processes. Apart from the duplex DNA, many other non-B DNA structures exist in cells such as hairpin, G-quadruplex, i-motif, triplex, cruciform, etc. There is a lack of information on the topology of these structures under torsional stress. My research work focused on addressing the population dynamics of non-B DNA tetraplex structures under torsionally constrained physiological conditions at the molecular level. For this study, we developed magneto-optical tweezers by combining dual-beam optical tweezers and the manipulation of magnetic tweezers for rotation.;The effect of torsional stress on the topology of non-B DNA tetraplexes was studied by varying the superhelicity of the template DNA through rotation of the magnets. For this study, we analyzed the sequence [d(ACAGGGGTGTGGGG) 2] from a promoter region of Insulin Linked Polymorphic Region and quantified the molecular population dynamics of DNA tetraplexes under various chemical (ions and pH) and mechanical (template superhelicity and molecular crowding) conditions by single-molecule mechanical unfolding methods. By mechanical unfolding of individual tetraplexes, we found that ions and pH have the most substantial effect on the formation of G-quadruplex and i-motif, respectively. Interestingly, superhelicity has the second largest effect followed by molecular crowding condition. While chemical effects are specific to tetraplex species mechanical factors show generic influences. This result provides substantial evidence for topology-based transcription modulation at the molecular level. Understanding the effect of torsional stress on the topology of secondary structures found in the chromosomes will serve as a deep insight into various cellular functions.
机译：最常见的B型脱氧核糖核酸（DNA）由两条由沃森-克里克碱基对（bp）连接在一起的聚合核酸链组成，每条10.5 bp处都有右旋。在细胞核中，DNA受到空间限制，因此会发生超螺旋（sigma），以通过过度缠绕（正超螺旋性），展开（负超螺旋性），DNA弯曲或熔化来缓解压力。在许多细胞过程中观察到的类似拓扑变化，例如转录，复制等，对这些过程的速率有重大影响。除了双链DNA以外，细胞中还存在许多其他非B DNA结构，例如发夹，G-四链体，i-基序，三链体，十字形等。在扭转应力下缺乏有关这些结构的拓扑的信息。我的研究工作集中于解决在分子水平上受扭转约束的生理条件下非B DNA四重体结构的种群动态。在这项研究中，我们通过结合双光束光学镊子和操纵磁性镊子旋转来开发了磁光镊子。;通过改变模板的超螺旋性研究了扭转应力对非B DNA四链体拓扑的影响DNA通过磁铁旋转。在这项研究中，我们分析了胰岛素连接多态性区域启动子区域的序列[d（ACAGGGGTGTGGGG）2]，并定量了在各种化学（离子和pH）和机械（模板超螺旋性和分子拥挤）下DNA四重体的分子种群动态。条件是通过单分子机械展开法。通过单个四链体的机械展开，我们发现离子和pH分别对G-四链体和i-基序的形成具有最重要的影响。有趣的是，超螺旋效应紧随其后的是分子拥挤状况。尽管化学效应对四重体物种特有，但机械因素却显示出一般性的影响。该结果为在分子水平上基于拓扑的转录调节提供了实质性证据。了解扭转应力对染色体中二级结构拓扑的影响，将有助于深入了解各种细胞功能。

著录项

  • 作者

    Selvam, Sangeetha.;

  • 作者单位

    Kent State University.;

  • 授予单位 Kent State University.;
  • 学科 Biochemistry.;Biophysics.;Chemistry.;Analytical chemistry.
  • 学位 Ph.D.
  • 年度 2018
  • 页码 137 p.
  • 总页数 137
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类
  • 关键词

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