Human apolipoprotein C-II forms twisted amyloid ribbons and closed loops

Human apolipoprotein C-II (apoC-II) self-associates in solution to form agg regates with the characteristics of amyloid including red-green birefringen ce in the presence of Congo Red under cross-polarized light, increased fluo rescence in the presence of thioflavin T, and a fibrous structure when exam ined by electron microscopy. ApoC-II was expressed and purified from Escher ichia 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 ap oC-II was initially soluble, eluting as low molecular weight species in gel filtration experiments using Sephadex G-50. Circular dichroism (CD) spectr oscopy indicated predominantly unordered structure. Upon incubation for 24 h, apoC-II self-associated into high molecular weight aggregates as indicat ed by elution in the void volume of a Sephadex G-50 column, by rapid sedime ntation in an analytical ultracentrifuge, and by increased light scattering . CD spectroscopy indicated an increase in beta-sheet content, while fluore scence emission spectroscopy of the single tryptophan revealed a blue shift and an increase in maximum intensity, suggesting repositioning of the tryp tophan into a less polar environment. Electron microscopy of apoC-II aggreg ates revealed a novel looped-ribbon morphology (width 12 nm) and several is olated closed loops. Like ail of the conserved plasma apolipoproteins, apoC -II contains amphipathic helical regions that account for the increase in c t-helix content on lipid binding. The increase in beta-structure accompanyi ng 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.