RUI: Single Molecule and Bulk Calorimetric Measurements of Insulin Binding Interactions with G-Quadruplex DNA

RUI：胰岛素与 G-四链体 DNA 结合相互作用的单分子和体量热测量

• 批准号: 1022022
• 负责人: Kumar Sinniah
• 金额: $ 21.19万
• 依托单位: Calvin University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-01 至 2014-12-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1022022&HistoricalAwards=false
• 关键词: RUI; Single; Molecule; Bulk; Calorimetric

Intellectual Merit: G-quadruplexes are noncanonical DNA structures formed from guanine-rich DNA sequences in the presence of monovalent cations such as potassium or sodium ions. These structures are of significant interest due to their possible role in biological processes. Since the human genome contains a large number of sequences that have the potential to form quadruplexes, proteins that bind to G-quadruplex DNA may provide a clue to the role of G-quadruplex DNA in biology. This project examines in detail the biophysical interactions between the protein insulin and G-quadruplex DNA. It has recently been reported that insulin binds with high selectivity to G-quadruplex DNA that contains the sequence of the insulin-linked polymorphic region (ILPR) of the human insulin promoter region. Given the probable role of the ILPR DNA sequence in gene regulation, the insulin binding to quadruplex DNA will be investigated by single molecule force spectroscopy and bulk ensemble techniques such as microcalorimetry. A key aspect of this research is to quantify the strength and the specificity of the binding interaction from single molecule to the bulk. The biophysical properties of G-quadruplex and the quadruplex binding proteins such as insulin are essential for understanding the biology of these important biomolecular systems. Broader Impacts: This research requires an interdisciplinary approach employing ideas and skills from biology, chemistry, physics and nanotechnology. A significant number of undergraduate students chosen from different departments at Calvin College, a primarily undergraduate institution, will work collaboratively gaining significant educational benefits from participating in a study that straddles several disciplines. This project will also provide excellent training in research and education in cross-disciplinary problem solving required by twenty-first century science. The research also provides students access to specialized instrumentation and research experiences that will enhance their analytical and creative thinking skills, while building confidence in their intellectual abilities. In addition to training undergraduate students, the knowledge gained from this research can also be applied to characterize other proteins that recognize G-quadruplexes. For example, proteins such as insulin-like growth factors are structurally similar to insulin, and they could be good candidates for determining whether their binding interaction is similar to insulin at the molecular level.

智力优点：G-四链体是非规范的 DNA 结构，由富含鸟嘌呤的 DNA 序列在单价阳离子（如钾或钠离子）存在下形成。 这些结构由于它们在生物过程中可能发挥的作用而引起人们的极大兴趣。 由于人类基因组包含大量有可能形成四链体的序列，因此与 G-四链体 DNA 结合的蛋白质可能为 G-四链体 DNA 在生物学中的作用提供线索。 该项目详细研究了胰岛素蛋白和 G-四链体 DNA 之间的生物物理相互作用。 最近有报道称，胰岛素高选择性地与含有人胰岛素启动子区胰岛素连接多态性区（ILPR）序列的G-四链体DNA结合。 考虑到 ILPR DNA 序列在基因调控中可能发挥的作用，胰岛素与四链体 DNA 的结合将通过单分子力谱和整体集成技术（例如微量热法）进行研究。 这项研究的一个关键方面是量化从单分子到本体的结合相互作用的强度和特异性。 G-四链体和四链体结合蛋白（例如胰岛素）的生物物理特性对于理解这些重要生物分子系统的生物学至关重要。 更广泛的影响：这项研究需要采用跨学科方法，运用生物学、化学、物理学和纳米技术的思想和技能。 从卡尔文学院（主要是本科院校）的不同院系中选出的大量本科生将通过参与跨多个学科的研究来合作，获得显着的教育效益。 该项目还将提供优秀的研究和教育培训，以解决二十一世纪科学所需的跨学科问题。该研究还为学生提供了专业仪器和研究经验，这将增强他们的分析和创造性思维技能，同时建立对他们智力能力的信心。 除了培训本科生外，从这项研究中获得的知识还可以应用于表征识别 G-四链体的其他蛋白质。 例如，胰岛素样生长因子等蛋白质在结构上与胰岛素相似，它们可能是确定它们的结合相互作用在分子水平上是否与胰岛素相似的良好候选者。