pH-Dependent Packing Mode Variations and Chirality Inversion in Short Peptide Self-Assembly

Precise control of structures and morphologies in peptide self-assembly has been challenging. We report the self-assembly of amphiphilic peptides I₃H, designed with a modular structure featuring three consecutive isoleucine residues as a hydrophobic tail and a C-terminal histidine-based hydrophilic headgroup. Microscopic, neutron scattering, and spectroscopic techniques demonstrate that the designed peptides self-assemble into β-sheet nanofibrils, with their helix handedness exhibiting subtle pH-dependent inversion. pH titration, NMR, and molecular dynamics simulations reveal the underlying mechanism correlates with the protonation state of histidine and the molecular packing modes in β-sheet assemblies. The protonated histidine promotes antiparallel β-sheet packing at lower pH while its deprotonated state favors parallel packing when pH is increased. Strong π-π stacking interactions between deprotonated histidine side chains in parallel β-sheet arrangements drive chiral flipping of β-strands, ultimately inducing supramolecular helix inversion. Furthermore, such a pH-dependent helix inversion can be engineered by inserting the achiral and flexible glycine at the hydrophobic/hydrophilic interface, with I₃GH assembly maintaining this effect while I₃GGH assembly abolishing it. This work not only advances our mechanistic understanding of peptide chirality inversion at the level of individual β-sheets but also provides a blueprint for designing hierarchical chirality through precise modulation of molecular packing modes and side-chain interactions.