title: pH-induced molecular shedding drives the formation of amyloid fibril-derived oligomers.
abstract: Amyloid disorders cause debilitating illnesses through the formation of toxic protein aggregates. The mechanisms of amyloid toxicity and the nature of species responsible for mediating cellular dysfunction remain unclear. Here, using β2-microglobulin (β2m) as a model system, we show that the disruption of membranes by amyloid fibrils is caused by the molecular shedding of membrane-active oligomers in a process that is dependent on pH. Using thioflavin T (ThT) fluorescence, NMR, EM and fluorescence correlation spectroscopy (FCS), we show that fibril disassembly at pH 6.4 results in the formation of nonnative spherical oligomers that disrupt synthetic membranes. By contrast, fibril dissociation at pH 7.4 results in the formation of nontoxic, native monomers. Chemical cross-linking or interaction with hsp70 increases the kinetic stability of fibrils and decreases their capacity to cause membrane disruption and cellular dysfunction. The results demonstrate how pH can modulate the deleterious effects of preformed amyloid aggregates and suggest why endocytic trafficking through acidic compartments may be a key factor in amyloid disease.
authors.affiliation: Astbury Centre for Structural Molecular Biology and School of Molecular and Cellular Biology, University of Leeds, Leeds, LS2 9JT, United Kingdom.
authors.forename: Kevin W
authors.initials: KW
authors.lastname: Tipping
authors.affiliation: Astbury Centre for Structural Molecular Biology and School of Molecular and Cellular Biology, University of Leeds, Leeds, LS2 9JT, United Kingdom.
authors.forename: Theodoros K
authors.initials: TK
authors.lastname: Karamanos
authors.affiliation: Astbury Centre for Structural Molecular Biology and School of Molecular and Cellular Biology, University of Leeds, Leeds, LS2 9JT, United Kingdom.
authors.forename: Toral
authors.initials: T
authors.lastname: Jakhria
authors.affiliation: Astbury Centre for Structural Molecular Biology and School of Molecular and Cellular Biology, University of Leeds, Leeds, LS2 9JT, United Kingdom.
authors.forename: Matthew G
authors.initials: MG
authors.lastname: Iadanza
authors.affiliation: Astbury Centre for Structural Molecular Biology and School of Molecular and Cellular Biology, University of Leeds, Leeds, LS2 9JT, United Kingdom.
authors.forename: Sophia C
authors.initials: SC
authors.lastname: Goodchild
authors.affiliation: Astbury Centre for Structural Molecular Biology and School of Molecular and Cellular Biology, University of Leeds, Leeds, LS2 9JT, United Kingdom.
authors.forename: Roman
authors.initials: R
authors.lastname: Tuma
authors.affiliation: Astbury Centre for Structural Molecular Biology and School of Molecular and Cellular Biology, University of Leeds, Leeds, LS2 9JT, United Kingdom.
authors.forename: Neil A
authors.initials: NA
authors.lastname: Ranson
authors.affiliation: Astbury Centre for Structural Molecular Biology and School of Molecular and Cellular Biology, University of Leeds, Leeds, LS2 9JT, United Kingdom.
authors.forename: Eric W
authors.initials: EW
authors.lastname: Hewitt
authors.affiliation: Astbury Centre for Structural Molecular Biology and School of Molecular and Cellular Biology, University of Leeds, Leeds, LS2 9JT, United Kingdom s.e.radford@leeds.ac.uk.
authors.forename: Sheena E
authors.initials: SE
authors.lastname: Radford
mesh_terms: D000682:Amyloid / Q000737:chemistry*; D000686:Amyloidosis / Q000378:metabolism; D052160:Benzothiazoles; D011992:Endosomes / Q000737:chemistry; D018840:HSP70 Heat-Shock Proteins / Q000737:chemistry; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008247:Lysosomes / Q000737:chemistry; D009000:Monocytes / Q000378:metabolism; D009113:Muramidase / Q000737:chemistry; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D013844:Thiazoles / Q000737:chemistry; D001613:beta 2-Microglobulin / Q000737:chemistry
chemical_list: D000682:Amyloid; D052160:Benzothiazoles; D018840:HSP70 Heat-Shock Proteins; D011994:Recombinant Proteins; D013844:Thiazoles; D001613:beta 2-Microglobulin; C009462:thioflavin T; D009113:Muramidase
keywords: amyloid; disassembly; fibrils; membrane disruption; oligomer