The Impact of Single Amino Acid Insertion on the Supramolecular Assembly Pathway of Aromatic Peptide Amphiphiles

Understanding the mechanism of self-assembly driven by non-covalent interactions is crucial for designing supramolecular materials with desired properties. Here we investigate the self-assembly of aromatic peptide amphiphiles, Fmoc-L₂QG and Fmoc-L₃QG using a combination of spectroscopic, transmission electron and superresolution optical microscopy techniques. Our results show that Fmoc-L₂QG leads to concentration-dependent assembly, forming fibrous assemblies at low concentrations and supramolecular droplets via liquid-liquid phase separation (LLPS) at higher concentrations. Mechanical activation using for example ultrasonication triggered the transition from metastable droplets to fibre morphologies of Fmoc-L₂QG. In contrast, Fmoc-L₃QG followed both on-pathway and off-pathway routes, resulting in the formation of fibrous morphologies regardless of concentration. Seeding experiments revealed that homo-seeds of the same peptide sequence accelerated the on-pathway process, while hetero-seeds of a mismatched peptide sequences accelerated the off-pathway process, highlighting the competing nature of the complex assembly profile. These findings demonstrate the significant impact of single amino acid insertion on the supramolecular assembly process of oligopeptide monomers, and highlight the potential for controlling the structure and dynamics of peptide materials. Pathway engineering of oligopeptide building blocks and multidomain supramolecular monomers will open new avenues in tailor-made and customizable supramolecular biomaterials.