{"id":{"repo_id":"lancaster","oai_identifier":"oai:eprints.lancs.ac.uk:84718"},"canonical_url":"https://search.dev.ndltd.org/etd/lancaster/oai:eprints.lancs.ac.uk:84718","repository":{"repo_id":"lancaster","name":"Lancaster University","base_url":"https://eprints.lancs.ac.uk/cgi/oai2"},"display":{"title":"Molecular level characterisation of apolipoprotein A-I aggregation leading to fibrils comprising of both α-helical and β-sheet structures","abstract":"Amyloidosis is defined as the misfolding of native proteins into insoluble fibrils that are deposited within tissues and extracellular organs. 30+ structurally and sequentially unrelated proteins have the ability to form amyloid aggregates, all of which contain characteristic features. ApoA-I, the main component in high-density lipoprotein, aggregates and becomes deposited as amyloid, either full-length apoA-I fibrils within atherosclerotic plaques, or N-terminal fragments of mutant apoA-I within organs. The work here aims to further the understanding of conditions that promote the aggregation of apoA-I in vitro, allowing the structural study of aggregated apoA-I at a molecular level. ApoA-I remains soluble at neutral pH, maintaining a predominantly α-helical conformation. Upon acidification to pH 4, apoA-I readily assembles into aggregates that, despite being responsive to the amyloid characteristic ThT dye, do not have the typical amyloid morphology and do not produce XRD diffraction patterns suggestive of β-sheets. The inclusion of heparin, and chemical oxidation of apoA-I methionine residues, in order to mimic physiological conditions, results in an increased ThT response and aggregated material more characteristic of amyloid. Solid-state NMR spectroscopy reveals for the first time that all three aggregation inducing conditions produce aggregates that give rise to cross-peaks corresponding to both α-helical and novel β-sheet structures. This leads to a refinement in the current theory describing apoA-I aggregation. In native apoA-I, the N-terminal 4-helical bundle protects the 3 hot spot regions from self-association into β-sheets. Acidification of apoA-I leads to the destabilisation of this N-terminus, and a conversion of residues 1-90 into β-sheet structures, whilst the C-terminus retains its α-helical structure. EGCG, an inhibitor of Aβ, α-synuclein and huntingtin amyloidosis, is shown here to bind to apoA-I with micro-molar affinity. However, rather than inhibit amyloidosis, EGCG causes a structural rearrangement of the aggregated material, resulting in a reduced α-helical content.","abstract_html":"Amyloidosis is defined as the misfolding of native proteins into insoluble fibrils that are deposited within tissues and extracellular organs. 30+ structurally and sequentially unrelated proteins have the ability to form amyloid aggregates, all of which contain characteristic features. ApoA-I, the main component in high-density lipoprotein, aggregates and becomes deposited as amyloid, either full-length apoA-I fibrils within atherosclerotic plaques, or N-terminal fragments of mutant apoA-I within organs. The work here aims to further the understanding of conditions that promote the aggregation of apoA-I in vitro, allowing the structural study of aggregated apoA-I at a molecular level. ApoA-I remains soluble at neutral pH, maintaining a predominantly α-helical conformation. Upon acidification to pH 4, apoA-I readily assembles into aggregates that, despite being responsive to the amyloid characteristic ThT dye, do not have the typical amyloid morphology and do not produce XRD diffraction patterns suggestive of β-sheets. The inclusion of heparin, and chemical oxidation of apoA-I methionine residues, in order to mimic physiological conditions, results in an increased ThT response and aggregated material more characteristic of amyloid. Solid-state NMR spectroscopy reveals for the first time that all three aggregation inducing conditions produce aggregates that give rise to cross-peaks corresponding to both α-helical and novel β-sheet structures. This leads to a refinement in the current theory describing apoA-I aggregation. In native apoA-I, the N-terminal 4-helical bundle protects the 3 hot spot regions from self-association into β-sheets. Acidification of apoA-I leads to the destabilisation of this N-terminus, and a conversion of residues 1-90 into β-sheet structures, whilst the C-terminus retains its α-helical structure. EGCG, an inhibitor of Aβ, α-synuclein and huntingtin amyloidosis, is shown here to bind to apoA-I with micro-molar affinity. However, rather than inhibit amyloidosis, EGCG causes a structural rearrangement of the aggregated material, resulting in a reduced α-helical content.","abstract_has_math":false,"creators":["Townsend, David John","Middleton, David"],"institution":"Lancaster University","degree_name":"Ph.D.","degree_level":"doctoral","degree_discipline":null,"degree_department":null,"school":null,"contributors":[],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2016,"date_issued":"2016","date_published":"2016","updated_at":"2026-07-24T02:48:32Z","subjects":[],"languages":[],"rights":[],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":null,"outbound_label":null,"outbound_source":null},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:creator","label":"Author","values":["Townsend, David John","Middleton, David"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["2016"]},{"key":"dc:date.issued","label":"Date","values":["2016"]},{"key":"dc:publisher.commercial","label":"Dc Publisher Commercial","values":["Lancaster University"]},{"key":"dc:publisher.institution","label":"Dc Publisher Institution","values":["Lancaster University"]},{"key":"dc:relation.isreferencedby","label":"Dc Relation Isreferencedby","values":["https://eprints.lancs.ac.uk/id/eprint/84718/"]},{"key":"dc:type","label":"Dc Type","values":["Thesis"]},{"key":"dc:type.qualificationlevel","label":"Dc Type Qualificationlevel","values":["doctoral"]},{"key":"dc:type.qualificationname","label":"Dc Type Qualificationname","values":["Ph.D."]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier.uri","label":"Identifier URI","values":["https://eprints.lancs.ac.uk/id/eprint/84718/1/2016TownsendPhD.pdf"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["Amyloidosis is defined as the misfolding of native proteins into insoluble fibrils that are deposited within tissues and extracellular organs. 30+ structurally and sequentially unrelated proteins have the ability to form amyloid aggregates, all of which contain characteristic features. ApoA-I, the main component in high-density lipoprotein, aggregates and becomes deposited as amyloid, either full-length apoA-I fibrils within atherosclerotic plaques, or N-terminal fragments of mutant apoA-I within organs. The work here aims to further the understanding of conditions that promote the aggregation of apoA-I in vitro, allowing the structural study of aggregated apoA-I at a molecular level. ApoA-I remains soluble at neutral pH, maintaining a predominantly α-helical conformation. Upon acidification to pH 4, apoA-I readily assembles into aggregates that, despite being responsive to the amyloid characteristic ThT dye, do not have the typical amyloid morphology and do not produce XRD diffraction patterns suggestive of β-sheets. The inclusion of heparin, and chemical oxidation of apoA-I methionine residues, in order to mimic physiological conditions, results in an increased ThT response and aggregated material more characteristic of amyloid. Solid-state NMR spectroscopy reveals for the first time that all three aggregation inducing conditions produce aggregates that give rise to cross-peaks corresponding to both α-helical and novel β-sheet structures. This leads to a refinement in the current theory describing apoA-I aggregation. In native apoA-I, the N-terminal 4-helical bundle protects the 3 hot spot regions from self-association into β-sheets. Acidification of apoA-I leads to the destabilisation of this N-terminus, and a conversion of residues 1-90 into β-sheet structures, whilst the C-terminus retains its α-helical structure. EGCG, an inhibitor of Aβ, α-synuclein and huntingtin amyloidosis, is shown here to bind to apoA-I with micro-molar affinity. However, rather than inhibit amyloidosis, EGCG causes a structural rearrangement of the aggregated material, resulting in a reduced α-helical content."]},{"key":"dc:format","label":"Dc Format","values":["application/pdf"]},{"key":"dc:title","label":"Title","values":["Molecular level characterisation of apolipoprotein A-I aggregation leading to fibrils comprising of both α-helical and β-sheet structures"]}]}],"canonical_facts":{"dc:creator":["Townsend, David John","Middleton, David"],"dc:date":["2016"],"dc:date.issued":["2016"],"dc:description.abstract":["Amyloidosis is defined as the misfolding of native proteins into insoluble fibrils that are deposited within tissues and extracellular organs. 30+ structurally and sequentially unrelated proteins have the ability to form amyloid aggregates, all of which contain characteristic features. ApoA-I, the main component in high-density lipoprotein, aggregates and becomes deposited as amyloid, either full-length apoA-I fibrils within atherosclerotic plaques, or N-terminal fragments of mutant apoA-I within organs. The work here aims to further the understanding of conditions that promote the aggregation of apoA-I in vitro, allowing the structural study of aggregated apoA-I at a molecular level. ApoA-I remains soluble at neutral pH, maintaining a predominantly α-helical conformation. Upon acidification to pH 4, apoA-I readily assembles into aggregates that, despite being responsive to the amyloid characteristic ThT dye, do not have the typical amyloid morphology and do not produce XRD diffraction patterns suggestive of β-sheets. The inclusion of heparin, and chemical oxidation of apoA-I methionine residues, in order to mimic physiological conditions, results in an increased ThT response and aggregated material more characteristic of amyloid. Solid-state NMR spectroscopy reveals for the first time that all three aggregation inducing conditions produce aggregates that give rise to cross-peaks corresponding to both α-helical and novel β-sheet structures. This leads to a refinement in the current theory describing apoA-I aggregation. In native apoA-I, the N-terminal 4-helical bundle protects the 3 hot spot regions from self-association into β-sheets. Acidification of apoA-I leads to the destabilisation of this N-terminus, and a conversion of residues 1-90 into β-sheet structures, whilst the C-terminus retains its α-helical structure. EGCG, an inhibitor of Aβ, α-synuclein and huntingtin amyloidosis, is shown here to bind to apoA-I with micro-molar affinity. However, rather than inhibit amyloidosis, EGCG causes a structural rearrangement of the aggregated material, resulting in a reduced α-helical content."],"dc:format":["application/pdf"],"dc:identifier.uri":["https://eprints.lancs.ac.uk/id/eprint/84718/1/2016TownsendPhD.pdf"],"dc:publisher.commercial":["Lancaster University"],"dc:publisher.institution":["Lancaster University"],"dc:relation.isreferencedby":["https://eprints.lancs.ac.uk/id/eprint/84718/"],"dc:title":["Molecular level characterisation of apolipoprotein A-I aggregation leading to fibrils comprising of both α-helical and β-sheet structures"],"dc:type":["Thesis"],"dc:type.qualificationlevel":["doctoral"],"dc:type.qualificationname":["Ph.D."]},"updated_at":"2026-07-24T02:48:32Z"}