Solvent-Modulated Energetics of Tetrameric DNA Polymorphism

我们建议继续我们的研究，以表征溶剂在调节生物聚合物的结合亲和力和构象偏好中所起作用的热力学含量。在之前的预算期间，我们的研究集中在研究水和水溶性共溶剂在蛋白质和核酸跃迁以及配体-蛋白质结合的能量学中的作用。同时，我们已经开始建立一个程序，在这个程序中，我们表征了溶剂和单个溶剂组分对DNA G-四链体和I-基序的能量学的影响。G-四链体和I-基序构成了一类具有重要生物学意义的非规范四聚体DNA结构。富含鸟嘌呤的DNA链容易折叠成各种G-四链结构，在这些G-四链结构中，鸟嘌呤碱基以稳定的氢键排列相互关联，即G-四联体。富含胞嘧啶的链在弱酸性到近中性的pH下有折叠成I基序构象的倾向。后者代表一种DNA构象，由两个平行的双链以反平行的方向插入彼此之间的四链排列。*G-四链体和I-基序在拓扑和结构上表现出极大的多样性，被认为具有生物学意义。核苷酸序列、稳定反离子的类型和环境条件(如渗透分子或乌鸦的存在)的细微变化导致G-四链体的拓扑结构发生了巨大的变化。在所要求的预算期间，我们建议探索水和水可混溶的助溶剂(如尿素和甘氨酸甜菜碱)以及各种阳离子的浓度和pH在指导非规范四链DNA基序多态趋势方面的作用。我们设计了一种实验策略，旨在开发一种预测算法，该算法可用于探索富含鸟嘌呤序列的构象偏好作为环境因素的函数及其富含胞嘧啶的互补链的存在。我们将使用差示扫描量热法、UV/Vis分光光度法、CD光谱、超声波测速仪和高精度密度测量。我们已经证明了这些实验方法的可行性和信息含量。*我们的长期目标是开发所需的预测算法，以评估自然产生的核酸中具有重要功能结构域的序列特定、结构特定和溶剂特定的构象偏好。随着在越来越多的生物体中成功地产生基因组序列，这样的能力变得非常可取。最终，人们想要评估基因组中偏爱特定结构基序的局部序列结构域是否对应于生物作用或控制的功能部位。

We propose to continue our studies to characterize the thermodynamic content of the role of solvent in modulating the binding affinity and conformational preferences of biopolymers. During the previous budget period, our studies were focussed on investigating the role of water and water-soluble cosolvents in the energetics of protein and nucleic acid transitions and ligand-protein binding. In parallel, we have begun to establish a program in which we characterize the influence of solvent and individual solvent components on the energetics of DNA G-quadruplexes and i-motifs. G-quadruplexes and i-motifs form an important class of noncanonical tetrameric DNA structures of biological importance. Guanine-rich DNA strands are prone to fold into various G-quadruplex structures in which guanine bases associate with each other in stable hydrogen-bonded arrangements, G-quartets. Cytosine-rich strands exhibit a propensity to fold into the i-motif conformation at slightly acidic to near neutral pH. The latter represents a DNA conformation with a tetraplex arrangement of two parallel-stranded duplexes intercalated into each other in an antiparallel orientation.******G-quadruplexes and i-motifs exhibit a great deal of diversity in topology and struc¬ture which is believed to be of biological relevance. Subtle changes in nucleotide sequence, type of the stabilizing counterion, and the environmental conditions (e. g., presence of osmolytes or crow¬ders) results in dramatic changes in the topology of a G-quadruplex. During the requested budget period, we propose to explore the role of water and water-miscible cosolvents (such as urea and glycine betaine) and the concentration of various cations and pH in guiding the polymorphic tendencies of noncanonical tetraplex DNA motifs. We have designed an experimental strategy aimed at developing a predictive algorithm that can be used to explore the conformational preferences of guanine-rich sequences as a function of environmental factors and the presence of their complementary cytosine-rich strands. We will employ differential scanning calorimetry, UV/Vis spectrophotometry, CD spectroscopy, ultrasonic velocimetry, and high precision densimetry. We already have demonstrated the feasibility and information content of these experimental methods.******Our long-term objective is to develop predictive algorithms needed to evaluate sequence-specific, structure-specific, and solvent-specific conformational preferences of functionally important domains within naturally occurring nucleic acids. With the success in genomic sequence generation in an increasingly large number of organisms, such a capacity is becoming highly desirable. Ultimately, one would like to assess if local sequence domains in the genome that favor particular structural motifs correspond to functional sites of biological action or control.**