Abstract/Summary

In peptides, alternating sequences of hydrophobic and hydrophilic residues in general promote β-sheet formation. On the other hand, liquid-liquid phase separation (LLPS) via self-coacervation is observed for certain peptides that can have a lower degree of sequence order due to specific intermolecular interactions (electrostatic, -stacking, cation-π and others). Here, we explore the interplay between fibril formation and LLPS in aqueous solution for tryptophan-arginine repeat peptides (WR)n with n=2-5. Peptide conformations are probed us-ing a combination of spectroscopic methods and x-ray diffraction while self-assembly is stud-ied through cryo-TEM, laser scanning confocal microscopy (LSCM), optical microscopy and small-angle X-ray scattering (SAXS). The pH-dependent aggregation is a particular focus. We find a general trend for extended nanotape structures to be favoured upon increasing the number of (WR) repeats. Coacervate droplets are observed due to LLPS for (WR)2 and (WR)3 at pH 12, whereas twisted nanotapes are formed by (WR)4 and (WR)5. Hydrogel for-mation is also observed at sufficiently high concentration at pH 12 for (WR)n with n=3-5. In contrast at lower pH, we find that (WR)2 forms micelles or globular aggregates whereas ex-tended β-sheet structures (fibrils or nanotapes) are observed for the longer sequences. The cytocompatibility of all five sequences was determined using an MTT assay on human dermal fibroblast cells, and this revealed that the peptides are well tolerated by these cells at low concentrations. Our results reveal remarkable pH-dependent phase behaviour of these model alternating peptides and we elucidate conditions for LLPS in short model (WR)2 and (WR)3 peptides.