title: NBD-labeled phospholipid accelerates apolipoprotein C-II amyloid fibril formation but is not incorporated into mature fibrils.
abstract: Human apolipoprotein (apo) C-II is one of several lipid-binding proteins that self-assemble into fibrils and accumulate in disease-related amyloid deposits. A general characteristic of these amyloid deposits is the presence of lipids, known to modulate individual steps in amyloid fibril formation. ApoC-II fibril formation is activated by submicellar phospholipids but inhibited by micellar lipids. We examined the mechanism for the activation by submicellar lipids using the fluorescently labeled, short-chain phospholipid 1-dodecyl-[(7-nitro-2-1,3-benzoxadiazol-4-yl)amino]-2-hydroxyglycero-3-phosphocholine (NBD-lyso-12-PC). Addition of submicellar NBD-lyso-12-PC increased the rate of fibril formation by apoC-II approximately 2-fold. Stopped flow kinetic analysis using fluorescence detection and low, non-fibril-forming concentrations of apoC-II indicated NBD-lyso-12-PC binds rapidly, on the millisecond time scale, followed by the slower formation of discrete apoC-II tetramers. Sedimentation velocity analysis showed NBD-lyso-12-PC binds to both apoC-II monomers and tetramers at approximately five sites per monomer with an average dissociation constant of approximately 10 μM. Mature apoC-II fibrils formed in the presence of NBD-lyso-12-PC were devoid of lipid, indicating a purely catalytic role for submicellar lipids in the activation of apoC-II fibril formation. These studies demonstrate the catalytic potential of small amphiphilic molecules in controlling protein folding and fibril assembly pathways.
authors: Department of Biochemistry and Molecular Biology, Bio21 Molecular Science and Biotechnology Institute, The University of Melbourne, Melbourne, Victoria 3010, Australia.
authors: Timothy M
authors: TM
authors: Ryan
authors: Michael D W
authors: MD
authors: Griffin
authors: Michael F
authors: MF
authors: Bailey
authors: Peter
authors: P
authors: Schuck
authors: Geoffrey J
authors: GJ
authors: Howlett
mesh_terms: D009327:4-Chloro-7-nitrobenzofurazan / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D000682:Amyloid / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D053304:Apolipoprotein C-II / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000378:metabolism; D001665:Binding Sites; D055162:Biocatalysis; D006801:Humans; D008823:Micelles; D010713:Phosphatidylcholines / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D015854:Up-Regulation / Q000502:physiology*
chemical_list: C552834:2-(6-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)hexanoyl-1-hexadecanoylglycero-3-phosphocholine; D000682:Amyloid; D053304:Apolipoprotein C-II; D008823:Micelles; D010713:Phosphatidylcholines; D009327:4-Chloro-7-nitrobenzofurazan