Understanding the Formation of Apoferritin Amyloid Fibrils

We present the optimization of experimental conditions to yield long, rigid apoferritin protein amyloid fibrils, as well as the corresponding fibrillation pathway. Fibril growth kinetics was followed using atomic force microscopy (AFM), transmission electron microscopy (TEM), dynamic light scattering (DLS), circular dichroism (CD), fourier-transform infrared spectroscopy (FTIR), and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Among the morphologies identified, we show that the conditions result in small aggregates, as well as medium and long fibrils. Extended incubation times led to progressive unfolding and hydrolysis of the proteins into very short peptide fragments. AFM, SDS-PAGE, and CD support a universal common fibrillation mechanism in which hydrolyzed fragments play the central role. These collective results provide convincing evidence that protein unfolding and complete hydrolysis of the proteins into very short peptide sequences are essential for the formation of the final apoferritin amyloid-like fibrils.

Automated summary

The study focused on optimizing experimental conditions to produce long and rigid apoferritin protein amyloid fibrils, and investigated the corresponding fibrillation pathway. Extended incubation times led to the progressive unfolding and hydrolysis of proteins into very short peptide fragments, which are essential for the formation of final apoferritin amyloid-like fibrils. AFM, SDS-PAGE, and CD supported a universal common fibrillation mechanism where hydrolyzed fragments played a central role.