Hydrogen exchange, proteins, folding, and misfolding

多年来，在这笔赠款的支持下开展的工作帮助发展了氢交换知识和 目前在蛋白质结构和功能研究中广泛采用的方法。在这笔赠款期间的进展 Period发现了一种基本的、但以前没有人怀疑的蛋白质属性。看起来蛋白质 由称为折叠的少量合作展开/重新折叠单元组成，并且这些单元 单元通过不同的中间形式形成并组装成一个路径序列，以逐步 构建天然蛋白质。本申请建议在细胞色素c中进一步检验这一假说。 此外，这一行为的一般性将用其他主要的蛋白质模型进行测试，使用葡萄球菌 首先是核酸酶，然后根据特定的标准选择其他模型蛋白。在其他工作中 继续发展氢交换知识，并将其用于理解蛋白质动力学 和信号传递，设计的突变微扰将被用于研究局部涨落机制 蛋白质和调节更远距离变化的内部蛋白质通路。所有这些工作都将使用 我们已经开发的特殊氢交换方法，将取决于核磁共振分析。相关 依靠质谱分析的氢交换实验，与其他方法一起， 将用于研究淀粉样蛋白结构及其前体形式。最初的重点将是α-突触核蛋白 (帕金森氏病)和A|3(阿尔茨海默病)，并将使用高通量方法 其他人，期待着药物发现的努力。 这里描述的研究旨在了解生物蛋白质机制的功能，无论是在 无论是健康还是疾病。这项工作已经发现并正在继续研究一种以前未被怀疑的 蛋白质分子的性质，它们的协作折叠子结构，这似乎解释了蛋白质是如何 折叠成它们的活动形式，以及它们如何执行其他一些功能。所获得的知识和特殊之处 开发的方法也被用于研究蛋白质错误折叠的机制，这些机制导致毁灭性的 人和动物的神经退行性病变，如帕金森氏症、阿尔茨海默病和牛 海绵状脑病(疯牛病)。

Work supported by this grant over the years helped to develop the hydrogen exchange knowledge and methods that are now widely exploited in studies of protein structure and function. Progress during this grant period has uncovered a fundamental but previously unsuspected protein property. It appears that proteins are composed of a small number of coooperative unfolding/refolding units, called foldons, and that these units form and assemble in a pathway sequence through distinct intermediate forms to progressively construct the native protein. The present application proposes to further test this hypothesis in cytochrome c. Also the generality of this behavor will be tested with other major protein models, using staphylococcal nuclease initially and then other model proteins to be selected according to specified criteria. In other work to continue the development of hydrogen exchange knowledge and its use for understanding protein dynamics and signal transfer, designed mutational perturbations will be used to study the local fluctuation mechanisms of proteins and the internal protein pathways that mediate more distant changes. All of this work will use special hydrogen exchange methods that we have developed and will depend on NMR analysis. Related hydrogen exchange experiments that depend on mass spectroscopic analysis, together with other methods, will be used to study amyloid structures and their precursor forms. The focus will be on a-synuclein initially (Parkinson's Disease) and then A|3 (Alzheimer's Disease) and will use high throughput methods among others, looking toward drug discovery efforts. The research described here aims to understand the functioning of the protein machinery of biology, both in health and in disease. This work has uncovered and is continuing to study a previously unsuspected property of protein molecules, their cooperative foldon substructure, which appears to explain how proteins fold to their active form and how they perform some other functions. The knowledge gained and the special methods developed are also being used to study the protein misfolding mechanisms that lead to devastating human and animal neurodegenerative pathologies such as Parkinson's and Alzheimer's disease and bovine spongiform encephalopathy (mad cow disease).