The transition state structure of protein aggregation

蛋白质聚集的过渡态结构

• 批准号: 345759759
• 负责人: Professor Dr. Alexander Buell
• 金额: --
• 依托单位: Institut für Physikalische Biologie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2017
• 资助国家: 德国
• 起止时间: 2016-12-31 至 2020-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/345759759?language=en
• 关键词: transition; state; structure; protein; aggregation

Proteins are the most versatile and complex class of all known molecules. Most biological processes crucially require one or more proteins in a very specific structural conformation and protein molecules often fold spontaneously into these structures. The mechanisms of protein folding have been studied in the last 20 years at the single amino acid residue level of detail, thanks to the advances of spectroscopic techniques and methodologies such as Phi-value analysis, a combination of accurate kinetic and thermodynamic measurements with protein engineering, allowing determination of the structures of the folding transition states. However, other than fold into their native state, proteins can also aggregate into ordered supramolecular structures, in particular into amyloid fibrils, and such processes are a hallmark of a range of severe neurodegenerative disorders, such as Alzheimer's and Parkinson's disease. Our fundamental mechanistic understanding of these aggregation processes lacks far behind that of protein folding. This lack of progress is partly due to the complex multistep nature of the aggregation process, which consists of nucleation, growth and potentially autocatalytic amplification steps, but also due to a lack of suitable experimental strategies to tackle this challenge. In this research project, I intend to develop a comprehensive approach to address this problem. The experimental strategy I propose consists of developing a variant of the Phi-value analysis (which has proven highly successful in the study of protein folding) which can be applied to protein misfolding and aggregation. This is in particular only possible for the growth step of the overall aggregation process. These experiments will be performed with a small protein domain (PI3K-SH3), because extensive data exists on the folding mechanism of SH3 domains, with which the data on the aggregation mechanism of this protein that will be obtained here, can be compared. The aim is hereby to gain structural information on the transition state, the state of highest energy between the monomeric and the aggregated protein. This will be the first experimental strategy to push the limits of mechanistic understanding of protein self-assembly and aggregation down to the level of individual amino acid residues.Furthermore, I aim to develop in this project a novel high throughput experimental strategy, with which it will be possible in the future to investigate a significantly larger number of sequence variant (hundreds, instead of dozens). The long term goal of this research program is to elucidate the exact aggregation mechanism of various proteins, in order to understand better the origin of this general phenomenon and in order to be able to control it.

蛋白质是所有已知分子中用途最多、结构最复杂的一类。大多数生物过程都需要一种或多种具有特定结构构象的蛋白质，蛋白质分子常常自发地折叠到这些结构中。在过去的20年里，由于光谱技术和方法学的进步，人们已经在单一氨基酸残基的详细水平上研究了蛋白质折叠的机制，如phi值分析，这是一种将精确的动力学和热力学测量与蛋白质工程相结合，从而能够确定折叠过渡态的结构。然而，除了折叠成自然状态外，蛋白质还可以聚集成有序的超分子结构，特别是淀粉样纤维，这样的过程是一系列严重神经退行性疾病的标志，如阿尔茨海默氏症和帕金森氏症。我们对这些聚集过程的基本机制的理解远远落后于蛋白质折叠。缺乏进展的部分原因是，聚集过程具有复杂的多步骤性质，包括成核、生长和潜在的自动催化放大步骤，但也由于缺乏适当的实验战略来应对这一挑战。在这个研究项目中，我打算开发一个全面的方法来解决这个问题。我提出的实验策略包括开发一种变种的phi-Value分析(它在蛋白质折叠研究中被证明非常成功)，可以应用于蛋白质的错误折叠和聚集。尤其是对于整个聚合过程的增长阶段，这是可能的。这些实验将用一个小的蛋白质结构域(PI3K-SH3)进行，因为关于SH3结构域的折叠机制有大量的数据，可以与这里获得的关于该蛋白质聚集机制的数据进行比较。因此，目的是获得关于过渡态的结构信息，过渡态是单体和聚集蛋白质之间的最高能量状态。这将是第一个将蛋白质自组装和聚集的机制理解的极限推向单个氨基酸残基水平的实验策略。此外，我的目标是在这个项目中开发一种新的高通量实验策略，它将有可能在未来研究更多数量的序列变体(数百个，而不是几十个)。这一研究计划的长期目标是阐明各种蛋白质的确切聚集机制，以便更好地了解这种普遍现象的起源，以便能够对其进行控制。

项目成果

Atomic structure of PI3-kinase SH3 amyloid fibrils by cryo-electron microscopy

• DOI: 10.1038/s41467-019-11320-8
• 发表时间: 2019-08-21
• 期刊: NATURE COMMUNICATIONS
• 影响因子: 16.6
• 作者: Roeder, Christine;Vettore, Nicola;Schroeder, Gunnar F.
• 通讯作者: Schroeder, Gunnar F.

Thermodynamics of amyloid fibril formation from non-equilibrium experiments of growth and dissociation.

生长和解离非平衡实验中淀粉样原纤维形成的热力学

• DOI: 10.1016/j.bpc.2021.106549
• 发表时间: 2021
• 期刊: Biophysical chemistry
• 影响因子: 3.8
• 作者: Rasmus K. Norrild;Nicola Vettore;Alberto Coden;Wei-Feng Xue;Alexander K. Buell
• 通讯作者: Alexander K. Buell