Single-Molecule Study of Biopolymers in Complex Solutions

ID: MPS/DMR/BMAT(7623) 1006737 PI: Saleh， Omar ORG：加州大学圣巴巴拉分校标题：复杂溶液中生物聚合物的单分子研究知识优势：该项目将研究不同序列的单链DNA （ssDNA）分子在各种溶液条件下的弹性，并量化多价反离子和拥挤剂对生物聚合物结构动态方面的影响。该研究计划的目的是在生物细胞中遇到的那种拥挤、富含盐的环境中，得出明确、定量的物理原理来控制离子生物聚合物的行为。PI将直接量化各种序列的ssDNA的物理参数，并试图解决关于ssDNA局部刚度序列依赖性的矛盾结果。他将研究ssDNA碱基堆叠创造棒状统计单体的能力；棒状聚合物已经被预测（虽然没有被证明）具有新的弹性行为。多价溶液静电是一个尚未解决的主要理论问题，对不同价盐的影响的研究将为解决溶液静电的复杂问题提供清晰的数据。这种方法将提供可能证实或否定某些理论的数据。分子拥挤是一种普遍的非特异性相互作用，它促进紧凑的生物分子构象，从而影响折叠生物聚合物的稳定性和大分子结合平衡。这些都是基本的和普遍存在的生物分子过程，提出的实验方法将为这个重要问题提供基本的见解。具体来说，PI将测量拥挤溶液中单个聚合物的构象，并将直接测试预测各种拥挤浓度下压实程度的最新理论。所有的实验都是为了帮助验证PI和同事最近在聚合物的缩放图和单分子拉伸实验之间建立的联系。提出的实验是基于PI实验室独特的低力拉伸能力。聚合物的低力弹性对单体之间的短期和长期相互作用都直接敏感，因此可以全面地探测聚合物的微观物理性质。在这方面，假设低力技术优于传统的高力方法，并且也可能比常用的散射技术具有优势。更广泛的影响：ssDNA的力学特性是各种生物、生物技术和生物材料问题的基础。包括非结构化ssdna在内的任何模型都必须准确地描述未折叠状态的序列依赖构象。该项目的数据将为这些模型提供基本的输入参数，从而影响我们对分子生物学过程（例如RNA折叠）和dsDNA的泡形成和变性转变的理解。此外，ssdna的力学响应的测量可以指导它们在基于dna的纳米结构中的合理使用和更好地设计分子信标。最后，这项工作的一部分将作为本科生实习生的暑期项目，这些实习生将从传统上在物理科学领域代表性不足的学生的校园项目中招募。实习生将直接与从事该项目的研究生一起工作。他们将学习各种各样适合这一跨学科工作的技能，包括生物分子合成、偶联化学、定量数据获取和分析。实习生们将参加小组会议，做演讲，并就他们的工作写一篇期末论文。因此，他们将获得开展尖端跨学科研究所需的广泛活动的大量接触和经验。最后，指导研究生将获得作为教师和研究项目主任的宝贵经验；这将直接为他未来作为一个研究小组负责人的科学生涯做好准备，同时通过更好地理解如何将日常研究与大局目标联系起来，提高他目前工作的质量。

ID: MPS/DMR/BMAT(7623) 1006737 PI: Saleh, Omar ORG: University of California, Santa BarbaraTitle: Single-Molecule Study of Biopolymers in Complex SolutionsINTELLECTUAL MERIT: The project will study the elasticity of single-stranded DNA (ssDNA) molecules of various sequences in a variety of solution conditions, and quantify the effects of multivalent counterions and crowding agents on dynamic aspects of biopolymer structure. The research program is designed to result in clear, quantitative physical principles governing ionic biopolymer behavior in the kind of crowded, salt-rich environments encountered in biological cells. The PI will directly quantify the physical parameters of ssDNA of various sequences, and attempt to resolve conflicting results on the sequence-dependence of ssDNA's local stiffness. He will investigate the ability of ssDNA base-stacking to create rod-like statistical monomers; rod-like polymers have been predicted (though not demonstrated) to have novel elasticity behaviors. Multivalent solution electrostatics represents a major unsolved theoretical problem, and this study of the effect of salts of various valencies will give clear data on complex issues of solution electrostatics. The approach will provide data that may validate or invalidate certain theories. Molecular crowding is a universal and non-specific interaction that promotes compact biomolecular conformations, thus affecting the stability of folded biopolymers and macromolecular binding equilibria. These are fundamental and ubiquitous biomolecular processes, and the proposed experimental approach to this problem will provide basic insight into this important problem. Specifically, the PI will measure the conformation of individual polymers in a crowded solution and will directly test recent theories that predict the magnitude of compaction at various crowded concentrations. All of the experiments are designed to help validate the link recently forged by the PI and coworkers between the scaling picture of polymers and single-molecule stretching experiments. The proposed experiments are based on the unique low-force stretching capabilities of the PI's lab. A polymer's low-force elasticity is directly sensitive to both the short- and long-range interactions between the monomers and thus can comprehensively probe the microscopic physics of the polymer. In this respect, the low-force technique is hypothesized to outperform traditional high-force methods, and may also have advantages over commonly-used scattering techniques.BROADER IMPACTS: The mechanical properties of ssDNA underlie a variety of biological, biotechnological, and biomaterials problems. Any model including unstructured ssDNAs must accurately describe the sequence-dependent conformation of the unfolded state. Data from this project will provide basic input parameters for such models, and thus will impact our understanding of processes in molecular biology (e.g. RNA folding) and bubble formation and denaturation transitions of dsDNA. Further, the measurements of the mechanical response of ssDNAs could lead to their rational use within DNA-based nanostructures and better design of molecular beacons. Finally, parts of this work will be used as a summer project for undergraduate interns, who will be recruited from on-campus programs for students traditionally under-represented in the physical sciences. The interns will work directly alongside the graduate student working on this project. They will learn a wide variety of skills appropriate to this interdisciplinary effort, including biomolecular synthesis, coupling chemistries, and quantitative data acquisition and analysis. The interns will attend group meetings, give presentations, and write a final paper on their work. Thus, they will gain significant exposure and experience to a wide range of activities needed to carry out cutting-edge interdisciplinary research. Finally, the mentoring graduate student will receive invaluable experience as a teacher and director of a research project; this should directly prepare him for a future scientific career as a research group leader, as well as improve the quality of his present work through a better understanding of how to relate day-to-day research with big-picture goals.