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dc.contributor.advisorGursky, Olgaen_US
dc.contributor.authorFrame, Nicholasen_US
dc.date.accessioned2019-11-22T16:25:22Z
dc.date.available2019-11-22T16:25:22Z
dc.date.issued2019
dc.identifier.urihttps://hdl.handle.net/2144/38578
dc.description.abstractSerum amyloid A (SAA) is a small, evolutionarily well-conserved, acute-phase protein best known as the protein precursor for amyloid A amyloidosis. During acute injury, infection, or inflammation, SAA plasma concentration rapidly rises 1000-fold, but the benefit of this dramatic increase is unclear. SAA functions in the innate immune response, cell signaling, and lipid homeostasis. Most SAA circulates on plasma high-density lipoproteins (HDL), where it reroutes HDL for lipid recycling. The aim of this dissertation is to provide a structural basis for understanding SAA-lipid interactions and to elucidate the structure-function relationship in this ancient protein. SAA is an intrinsically disordered protein that acquires ~50% helical structure when bound to lipids, and is ~80% helical in three available atomic-resolution x-ray crystal structures. We took advantage of these crystal structures of lipid-free SAA to propose the binding site for various lipids, including lipids in HDL. We postulated that SAA, as a monomer, binds lipids via two amphipathic helices, h1 and h3, that form a concave hydrophobic surface, and that the curvature of this surface defines the binding preference of SAA for HDL versus larger lipoproteins. Next, we used murine SAA1.1 and a membrane-mimicking model phospholipid, palmitoyl-oleoyl phosphocholine (POPC), to reconstitute SAA-lipid complexes and characterize their overall structure, stability and stoichiometry using an array of spectroscopic, electron microscopic, and biochemical methods. We observed preferential formation of ~10 nm particles that mimic HDL size, accompanied by the α-helical folding. To probe the local protein conformation and dynamics in these SAA-POPC particles, we used hydrogen-deuterium exchange mass spectrometry. Analysis of the amount and the kinetics of deuterium uptake clearly established h1 and h3 as the lipid-binding site. Moreover, we determined that SAA binding to lipid follows a mixed model that combines induced fit, promoting α-folding in h3, with conformational selection, stabilizing pre-existing conformations in h1 and around the h2-h3 linker. Taken together, our results provided the structural basis necessary for understanding SAA-lipid interactions, which are central to beneficial functions of SAA as a housekeeping molecule, and to its misfolding in amyloid. This research sets the stage for understanding SAA interactions with its numerous other functional ligands.en_US
dc.language.isoen_US
dc.subjectBiophysicsen_US
dc.subjectIntrinsically disordered proteinen_US
dc.subjectLipoprotein nanoparticleen_US
dc.subjectProtein-lipid bindingen_US
dc.subjectProtein structureen_US
dc.titleThe structural basis for lipid interactions of serum amyloid Aen_US
dc.typeThesis/Dissertationen_US
dc.date.updated2019-10-07T19:02:11Z
etd.degree.nameDoctor of Philosophyen_US
etd.degree.leveldoctoralen_US
etd.degree.disciplineBiophysicsen_US
etd.degree.grantorBoston Universityen_US
dc.identifier.orcid0000-0003-0392-2900


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