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<b>Understanding the folding of amyloids using cryo-EM: </b><b><i>In vitro </i></b><b>studies and methods development</b>

<p dir="ltr">Neurodegenerative diseases are progressive, incurable conditions that affect tens of millions of people worldwide and are characterized by the aggregation of misfolded protein in the brain. Though the precise role of these amyloid aggregates in the onset and progression of these diseases is not clear at this time, there is a pressing need to understand how they form and spread in human disease. In service to these aims, I have conducted three small projects to expand knowledge in this regard. I first investigated the use of thioflavin T, a common amyloid stain, as an affinity reagent for the general purification of amyloid filaments from <i>ex vivo </i>samples, observing strong potential using a relatively simple, inexpensive magnetic bead conjugation technique. I next analyzed the formation of filaments of a truncated recombinant amyloid-beta peptide with residues 1-35, observing a new filament type formed at low pH in the wild-type sequence of this truncated peptide. Finally, I conducted structural studies on amyloid-beta(1-42) filaments prepared under different conditions consistent with traumatic brain injury to observe their effect on amyloid folding. While I found no effect of differential conditions on filament type, the low-resolution structures solved were highly consistent with aggregates found in Alzheimer’s disease patients, presenting a promising way forward for <i>in vitro</i> modeling of amyloid filaments that are true to pathology. In sum, the work here presented advances the concepts of both how amyloid aggregates from patient brains can be best prepared for structural analysis, and the factors underpinning their aggregation at the onset of neurodegenerative disease.</p>

  1. 10.25394/pgs.25647885.v2
Identiferoai:union.ndltd.org:purdue.edu/oai:figshare.com:article/25647885
Date18 April 2024
CreatorsRyan Patrick Kreiser (18405978)
Source SetsPurdue University
Detected LanguageEnglish
TypeText, Thesis
RightsCC BY 4.0
Relationhttps://figshare.com/articles/thesis/_b_Understanding_the_folding_of_amyloids_using_cryo-EM_b_b_i_In_vitro_i_b_b_studies_and_methods_development_b_/25647885

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