Fig. 5.

Characterization of ApoE4 fibrils. (a) Transmission electron micrographs of negatively stained ApoE4 (25 μM in 20 mM PB, pH 7.4) monitored over 24 h at 37 °C. The scale bar represents 200 nm. ApoE4 self-assembly was characterized by measuring changes in length (b) and width (c) of the fibrils. Changes in width were non-significant; however, with increasing incubation times, ApoE4 fibrils become significantly longer (average length of 363 nm; one-way ANOVA: ****p < 0.0001, F = 180.9). (d) CD spectra show retention of α-helical secondary structure after assembly (whole fraction). Fibrils in the pellet were separated from the supernatant to confirm that their secondary structure is not masked by protein in the supernatant. Fibrils in the pellet showed an α-helical conformation. (e) X-ray fiber diffraction pattern obtained from partially aligned ApoE4 fibrils after 24-h incubation at 37 °C showing positions of diffraction signals on the meridian (vertical) and equatorial (horizontal) axes.