Affinity purification of cross-ß fibrils using immobilized thioflavin

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Project Summary Neurodegenerative diseases such as Alzheimer’s disease cause a significant socioeconomic burden. These diseases manifest as protein plaques in the brain tissues. As of today, there are no available drugs to treat or prevent these diseases. For rational drug design, it is essential to understand the atomic details of the proteins involved in these plaques. To achieve this, researchers utilize structural biology methods such as electron cryomicroscopy (cryoEM), nuclear magnetic resonance, and X-ray crystallography. Neuropathologists isolate these plaques from post-mortem patient brain tissues and assess the disease for the presence of plaques. Then the samples are subjected to structural studies. Since 2017, a significant number of atomic structures of the protein molecules involved in the plaques have been reported. However, isolation of proteins from tissue samples is a bottleneck due to low yield, differences in the protein structures, and the presence of contaminants. The proteins in the plaques have a distinct arrangement in that they form a β-strand conformation and stack on top of each other along a helical axis. This arrangement gives rise to a unique property – they bind to molecules such as thioflavin and exhibit characteristic fluorescence emission. Such a property enables researchers to know the presence of protein plaques and quantify them in cellular models. Here, we propose to utilize the specificity of thioflavin to protein filaments of neuropathological plaques as a new method of plaque purification. The method combines the specificity of thioflavin to the protein filaments and affinity-based purification methods that are widely used to purify proteins. We propose that our method would enable researchers to extract and isolate protein filaments from brain tissues with much improved yields, fewer contaminants, and shorter processing times.