Abstract / Description of output
Human apolipoprotein C-II (apoC-II) self-associates in solution to form aggregates with the characteristics of amyloid including red-green birefringence in the presence of Congo Red under cross-polarized light, increased fluorescence in the presence of thioflavin T, and a fibrous structure when examined by electron microscopy. ApoC-II was expressed and purified from Escherichia coli and rapidly exchanged from 5 M guanidine hydrochloride into 100 mM sodium phosphate, pH 7.4, to a final concentration of 0.3 mg/mL. This apoC-II was initially soluble, eluting as low molecular weight species in gel filtration experiments using Sephadex G-50. Circular dichroism (CD) spectroscopy indicated predominantly unordered structure. Upon incubation for 24 h, apoC-II self-associated into high molecular weight aggregates as indicated by elution in the void volume of a Sephadex G-50 column, by rapid sedimentation in an analytical ultracentrifuge, and by increased light scattering. CD spectroscopy indicated an increase in beta-sheet content, while fluorescence emission spectroscopy of the single tryptophan revealed a blue shift and an increase in maximum intensity, suggesting repositioning of the tryptophan into a less polar environment. Electron microscopy of apoC-II aggregates revealed a novel looped-ribbon morphology (width 12 nm) and several isolated closed loops. Like ail of the conserved plasma apolipoproteins, apoC-II contains amphipathic helical regions that account for the increase in ct-helix content on lipid binding. The increase in beta-structure accompanying apoC-II fibril formation paints to an alternative folding pathway and an in vitro system to explore the general tendency of apolipoproteins to form amyloid in vivo.
Original language | English |
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Pages (from-to) | 8276-8283 |
Number of pages | 8 |
Journal | Biochemistry |
Volume | 39 |
Issue number | 28 |
DOIs | |
Publication status | Published - 18 Jul 2000 |
Keywords / Materials (for Non-textual outputs)
- LOW-DENSITY LIPOPROTEINS
- X-RAY-DIFFRACTION
- ALZHEIMERS-DISEASE
- HUMAN-PLASMA
- A-II
- PROTEIN
- PEPTIDES
- FIBRILS
- LIPASE
- AGGREGATION