KINETICS OF EARLY EVENTS IN PROTEIN FOLDING

蛋白质折叠早期事件的动力学

• 批准号: 2910355
• 负责人: HEINRICH RODER
• 金额: $ 20.61万
• 依托单位: RESEARCH INST OF FOX CHASE CAN CTR
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-05-01 至 2002-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2910355
• 关键词: Staphylococcus; chemical kinetics; cytochrome c; nuclease; protein biosynthesis; protein folding; structural biology; ubiquitin

DESCRIPTION: Despite an astronomical number of possible conformations, most naturally evolved polypeptides have acquired the ability to spontaneously fold into a unique three-dimensional structure, which gives rise to a protein's biological activity. A detailed structural, energetic and kinetic description of the reversible folding process in vitro for a representative set of proteins is an essential step toward understanding the underlying protein folding in the cell, and its medical implications. Because of the limited time resolution of conventional kinetic methods, little is known about the process of folding on the sub-millisecond time scale. These early structural events are the key to understanding how protein folding is initiated and are the main focus of many current theoretical models. Thus, the primary objective of this project is to directly observe the formation of partially folded early intermediates known to appear for many proteins during the first few milliseconds of refolding. A recently developed and extensively tested capillary mixing device makes it possible to extend the time resolution of continuous-flow fluorescence and quenched-flow hydrogen exchange studies into the microsecond time scale. This approach will be used to study the dynamics of early structural events, such as the initial chain collapse, formation of stable hydrogen bonds and long-range tertiary interactions for a series of well-characterized proteins and some mutants, with engineered fluorescence probes. Among these are mitochondrial and bacterial cytochromes c, ubiquitin and staphylococcal nuclease, which have been characterized extensively in Dr. Roder's laboratory by conventional structural and kinetic methods. The results will provide unique insight into the structural and dynamic properties of early structural intermediates and their kinetic role in protein folding.

描述：尽管可能的构象有天文数字，但大多数 自然进化的多肽已经获得了自发地 折叠成独特的三维结构，从而产生 蛋白质的生物活性。 详细的结构、能量和动力学 代表性的体外可逆折叠过程的描述 一组蛋白质是了解其潜在机制的重要一步 细胞中的蛋白质折叠及其医学意义。 因为 传统动力学方法的时间分辨率有限，人们知之甚少 关于亚毫秒时间尺度上的折叠过程。 这些早 结构事件是理解蛋白质折叠的关键 发起并是许多当前理论模型的主要焦点。 因此， 该项目的主要目标是直接观察地层 已知出现在许多蛋白质中的部分折叠的早期中间体 在重折叠的最初几毫秒内。 最近开发和 经过广泛测试的毛细管混合装置可以延长 连续流荧光和猝灭流氢的时间分辨率 交流微秒时间尺度的研究。 这种方法将 用于研究早期结构事件的动力学，例如初始 链塌陷，形成稳定的氢键和长程叔键 一系列特征明确的蛋白质和一些突变体的相互作用， 与工程荧光探针。 其中包括线粒体和 细菌细胞色素 c、泛素和葡萄球菌核酸酶，它们具有 Roder 博士的实验室通过常规方法进行了广泛的表征 结构和动力学方法。 结果将提供独特的见解 研究早期结构中间体的结构和动态特性 及其在蛋白质折叠中的动力学作用。