How do proteins fold: Mechanism and principles

这个项目将研究驱动蛋白质折叠的机制和原理。蛋白质折叠是所有生物的基础。将蛋白质从最初合成的非结构化状态运送到其生物功能结构的中间结构的途径可以说是所有生物学中最重要的反应。此外，它是理解许多正常过程和努力阻止具有挑战性的错误折叠过程的关键。折叠问题已被证明是极其困难的。所有可用的方法都针对这个问题，但它们通常不能描述动力学折叠过程中详细的结构形成。该实验室最近开发并展示了一种名为氢交换质谱仪(HX-MS)的新方法，它为深入研究甚至可能解开这个长期存在的谜团展示了巨大的希望。对折叠过程的了解可以提供必要的知识，以便以建设性的方式干扰错误的折叠过程。该调查员在培训研究生和博士后方面有着出色的记录，并将继续这些培训活动。此外，拟议的方法开发和软件工具可能使整个科学界受益。本实验室最近的工作发展了脉冲标记HX MS方法，并首次表明它可以用于定义共同构建蛋白质折叠途径的日益折叠的中间形式的结构。到目前为止，对三种蛋白质(核糖核酸酶H、细胞色素c和麦芽糖结合蛋白)的结果清楚地显示了将最初未折叠的蛋白质(U)通过中间体(II)运送到最终天然状态(N)的结构的定时发展。此外，研究结果还提出了三个似乎支配折叠过程的原则。未来的计划是测试这些结果的真实性和普遍性。最初的努力将针对色氨酸合成酶的阿尔法亚单位，这是一种中等大小的蛋白质，在TIM桶结构中有265个残基，代表最常见的蛋白质折叠，约占所有已知蛋白质的10%。然后，这项研究将扩展到小的全β蛋白泛素(76个残基)，然后最终扩展到巨大的Hsp104异构体(908个残基)。这一裁决反映了NSF的法定使命，并通过使用基金会的智力价值和更广泛的影响审查标准进行评估，被认为值得支持。

This project will investigate the mechanisms and principles that drive protein folding. Protein folding is fundamental to all living organisms. The pathway of intermediate structures that carry proteins from their initially synthesized unstructured condition to their biologically functional structure is arguably the most centrally important reaction in all of biology. Further, it is key to understanding many normal processes and to the effort to interdict challenging misfolding processes. The folding problem has proven extremely difficult. All available methodologies have been directed at the problem but they commonly cannot describe detailed structure formation during the kinetic folding process. This laboratory has developed and recently demonstrated a new approach, named hydrogen exchange analyzed by mass spectrometry (HX-MS), that shows great promise for delving into and perhaps solving this long-lasting enigma. The understanding of the folding process may provide the knowledge necessary to interfere in a constructive way with misfolding processes. The investigator has a superb record of training graduate students and postdocs and will continue these training activities. In addition, the proposed method development and software tools are likely to benefit the scientific community at large. Recent work in this laboratory has developed the pulse labeling HX MS method and shown it can be used to define, for the first time, the structure of the increasingly folded intermediate forms that together construct the protein folding pathway. Results so far obtained for three proteins (ribonuclease H, cytochrome c, and maltose binding protein) clearly display the timed development of structures that carry the initially unfolded proteins (U) through intermediate forms (Ii) to their final native state (N). Further the results suggest three principles that appear to govern the folding process. The plan going forward is to test for the reality and generality of these results. Initial efforts will be directed at the alpha-subunit of tryptophan synthase, a moderate-sized protein with 265 residues in a TIM barrel structure, which represents the most common protein fold, accounting for ~10% of all known proteins. The investigation will then be extended to the small all-beta protein ubiquitin (76 residues) and then ultimately to the immense Hsp104 protomer (908 residues).This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.