Liquid-Liquid Phase Separation (LLPS)-Driven Fibrilization of Amyloid- Protein

Amyloid-beta [A beta(1-40)] aggregation into a fibrillar network is one of the major hallmarks of Alzheimer's disease (AD). Recently, a few studies reported that polyphosphate (polyP), an anionic biopolymer that participates in various cellular physiological processes in humans, induces fibrilization in many amyloidogenic proteins [ -; and -]. However, the role of polyP in A beta(1-40) fibrilization and the underlying mechanism are unclear. In this study, we report experimental investigations on the role of polyP in the fibrilization kinetics of A beta(1-40). It is found that polyP exhibits a dual effect depending upon the pH value. At pH = 7 (neutral), polyP inhibits amyloid fibrilization in a dose-dependent manner similar to negatively charged nanoparticles. On the contrary, at pH = 3 (acidic), polyP accelerates amyloid fibrilization kinetics via liquid-liquid phase separation (LLPS), wherein the protein-rich droplets contain mature fibrils. In the parameter space spanned by concentrations of A beta(1-40) and polyP, a phase diagram is constructed to demark the domain where LLPS is observed at pH = 3. Characterization of the protein aggregates, secondary structure content in the aggregates, and cell viability studies in the presence of aggregates are discussed at both pH values. This study reveals that anionic biopolymers can modulate amyloid fibrilization kinetics, linked to neurodegenerative diseases, depending upon their local concentrations and pH.