Driving liquid-to-solid transitions in peptide-based polyelectrolyte complexes

Membraneless organelles (MLOs) are biological condensates in the body that are involved in essential cellular functions such as compartmentalization and catalysis of intercellular chemical reactions. Liquid-to-solid transitions of membraneless organelles have been linked to many diseases such as amyotrophic lateral sclerosis, Alzheimer’s disease, Creutzfeldt-Jakob disease and various cancers. Material properties of membraneless organelles include liquid-like properties as well as formation via liquid-liquid phase separation. Polyelectrolyte complexes (PECs), formed upon mixing a polycation and polyanion, also form via liquid-liquid phase separation and can form liquid droplets (coacervates) or solid aggregates. Using a peptide-based polyelectrolyte system allows for fine-tuning of parameters such as cation-pi interactions and charge density, which are also found in membraneless organelles. Manipulating the chiral patterning of peptides allows for the formation of both liquid and solid PECs. In this study, a library of peptide-based PECs was used to investigate liquid to solid transitions of coacervates. Upon addition of a solid complex into a solution of liquid coacervate complex, a liquid to solid transition was observed. Transitions were investigated using optical microscopy as well as fluorescence microscopy with Thioflavin-T, a dye that fluoresces in the presence of beta-sheet secondary structure formation. The percent change in secondary structure of liquid droplets as they transitioned into solid aggregates upon the addition of solid PECs was quantified through deconvolution analysis of FT-IR spectra in the amide I region (1600-1700 cm^-1).  Results showed that the percentage of beta-sheet secondary structure and type of beta-sheet secondary structure of solid-PECs did not play a role in their ability to generate liquid to solid transitions. On the other hand, solid PECs with greater hydrophobicity were able to transition liquid coacervate droplets at lower concentrations. Increasing the chain length of peptides was also shown to increase the ability of solid complexes to generate a liquid to solid transition. Interactions between peptides in the mixed system containing both liquid and solid PECs were further investigated with turbidimetric studies probing the critical salt concentration of new complexes formed with the liquid anion and solid cation and the solid anion and liquid cation in order to determine the mechanism of transition. Results illustrated that there was not a clear correlation between the critical salt of the new complex pairs and the ability of a solid complex to transition liquid droplets.