Structural Characterization and Amyloidogenic Pathways of Lipid-free and Lipid-bound Serum Amyloid A

SAA is an apolipoprotein that gets overexpressed by a 1000-fold during the acute phase response, making it a useful clinical biomarker of inflammation. SAA is normally bound to high-density lipoprotein (HDL) in the serum and helps orchestrate the innate immune response and affects lipid transport. In patients with long-term inflammation such as rheumatoid arthritis, obesity, Crohn’s disease, or long-covid, misfolding and fibrilization of SAA becomes a major risk. Misfolding of SAA causes AA Amyloidosis, a secondary disease state in which organ failure is caused by the extracellular deposition of cytotoxic fibrils that are composed of fragments or intact SAA protein. The molecular mechanisms and in vivo conditions that trigger SAA’s misfolding from an alpha helical to a beta sheet amyloid assembly during chronic inflammation are largely unknown. The structure of SAA in the HDL-bound form or the lipid-free form has never been studied in solution. This thesis aims to identify key structural features between lipid-bound and lipid-free SAA and to examine the conditions that trigger SAA Amyloidosis. First, the structure of SAA was studied in solution for the first time using techniques such as SAXS, TEM, CD, and SEC-MALS. A novel octameric lipid-free SAA was found that has similar dimensions as lipid-bound SAA nanodiscs. The solution structures of SAA allow us to better understand the formation of the lipid-bound state and suggests that relatively few conformational rearrangements may be required to allow SAA interactions with lipid-containing particles such as HDL. Next, two novel conditions were discovered which led to SAA amyloids: 1) interaction of lipid-bound SAA with physiologically relevant concentrations of calcium cations and 2) binding of an amyloidogenic peptide 54SFYVYSRVK62 or SK9 from SARS-CoV-2 envelope protein. These results highlight the importance of lipid interaction with the amyloidogenic core sequence of SAA in preventing it from self-assembly into amyloid. Factors such as viral peptides containing amyloidogenic core sequences or lipid-cation interactions can lead to the destabilization of the SAA native fold, promoting misfolding and subsequent formation of proto-fibrillar oligomers and amyloid fibrils. Understanding the structural dynamics and triggers of SAA misfolding into amyloid fibrils sheds light on potential therapeutic targets for preventing SAA amyloidosis in chronic inflammatory conditions, offering new avenues for intervention in related diseases.

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