KINETICS OF EARLY EVENTS IN PROTEIN FOLDING

描述：尽管有天文数字的可能构象，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， 蛋白质的生物活性。 详细的结构，能量和动力 代表性的体外可逆折叠过程的描述 一组蛋白质是了解潜在的 细胞中的蛋白质折叠及其医学意义。 因为 由于传统动力学方法的时间分辨率有限， 折叠的过程是亚毫秒级的 这些早期 结构事件是理解蛋白质折叠的关键 它是当前许多理论模型的主要焦点。 因此，在本发明中， 该项目的主要目的是直接观察 许多蛋白质的部分折叠的早期中间体 在重折叠的最初几毫秒。 最近开发的一种 经过广泛测试的毛细管混合装置使延长 连续流荧光和猝灭流氢的时间分辨 交流研究到微秒时间尺度。 这种方法将是 用于研究早期结构事件的动力学，例如最初的 链崩溃，形成稳定的氢键和长程三级 一系列特征明确的蛋白质和一些突变体的相互作用， 用工程荧光探针。 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， 在罗德博士的实验室中， 结构和动力学方法。 结果将提供独特的见解 早期结构中间体的结构和动力学性质 以及它们在蛋白质折叠中的动力学作用。

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.