title: A structural core within apolipoprotein C-II amyloid fibrils identified using hydrogen exchange and proteolysis.
abstract: Plasma apolipoproteins show alpha-helical structure in the lipid-bound state and limited conformational stability in the absence of lipid. This structural instability of lipid-free apolipoproteins may account for the high propensity of apolipoproteins to aggregate and accumulate in disease-related amyloid deposits. Here, we explore the properties of amyloid fibrils formed by apolipoproteins using human apolipoprotein (apo) C-II as a model system. Hydrogen-deuterium exchange and NMR spectroscopy of apoC-II fibrils revealed core regions between residues 19-37 and 57-74 with reduced amide proton exchange rates compared to monomeric apoC-II. The C-terminal core region was also identified by partial proteolysis of apoC-II amyloid fibrils using endoproteinase GluC and proteinase K. Complete tryptic hydrolysis of apoC-II fibrils followed by centrifugation yielded a single peptide in the pellet fraction identified using mass spectrometry as apoC-II(56-76). Synthetic apoC-II(56-76) readily formed fibrils, albeit with a different morphology and thioflavinT fluorescence yield compared to full-length apoC-II. Studies with smaller peptides narrowed this fibril-forming core to a region within residues 60-70. We postulate that the ability of apoC-II(60-70) to independently form amyloid fibrils drives fibril formation by apoC-II. These specific amyloid-forming regions within apolipoproteins may underlie the propensity of apolipoproteins and their peptide derivatives to accumulate in amyloid deposits in vivo.
authors.affiliation: Department of Biochemistry and Molecular Biology, Bio21 Molecular Science and Biotechnology Institute, The University of Melbourne, Parkville, Victoria 3010, Australia.
authors.forename: Leanne M
authors.initials: LM
authors.lastname: Wilson
authors.forename: Yee-Foong
authors.initials: YF
authors.lastname: Mok
authors.forename: Katrina J
authors.initials: KJ
authors.lastname: Binger
authors.forename: Michael D W
authors.initials: MD
authors.lastname: Griffin
authors.forename: Haydyn D T
authors.initials: HD
authors.lastname: Mertens
authors.forename: Feng
authors.initials: F
authors.lastname: Lin
authors.forename: John D
authors.initials: JD
authors.lastname: Wade
authors.forename: Paul R
authors.initials: PR
authors.lastname: Gooley
authors.forename: Geoffrey J
authors.initials: GJ
authors.lastname: Howlett
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000682:Amyloid / Q000737:chemistry* / Q000648:ultrastructure; D053304:Apolipoprotein C-II / Q000737:chemistry* / Q000302:isolation & purification / Q000648:ultrastructure; D052160:Benzothiazoles; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D003903:Deuterium / Q000378:metabolism*; D041961:Deuterium Exchange Measurement; D004305:Dose-Response Relationship, Drug; D004587:Electrophoresis, Agar Gel; D019286:Endopeptidase K / Q000494:pharmacology; D005456:Fluorescent Dyes; D006801:Humans; D006859:Hydrogen / Q000378:metabolism*; D006868:Hydrolysis; D007700:Kinetics; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D011487:Protein Conformation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D012697:Serine Endopeptidases / Q000494:pharmacology; D013050:Spectrometry, Fluorescence; D013844:Thiazoles; D013997:Time Factors; D014357:Trypsin / Q000494:pharmacology
chemical_list: D000682:Amyloid; D053304:Apolipoprotein C-II; D052160:Benzothiazoles; D005456:Fluorescent Dyes; D013844:Thiazoles; C054300:apolipoprotein C-II (St Michael); C009462:thioflavin T; D006859:Hydrogen; D003903:Deuterium; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase; D014357:Trypsin; D019286:Endopeptidase K