Halogen-Controlled Aromatic Interactions Drive the Self-Assembly and Mechanics of Peptide Hydrogels

-halogen substitution (F, Cl, Br, I) on the phenylalanine residue of the amyloidogenic hexapeptide FINyVK, derived from the C-terminal domain of nucleophosmin 1 (NPM1). A systematic combination of spectroscopic, rheological, morphological, and biological analyses was employed to elucidate how halogen identity influences peptide aggregation and hydrogel formation across multiple length scales. All halogenated analogues exhibited a marked decrease in critical aggregation concentration (CAC) with increasing halogen size, reflecting enhanced hydrophobic and aromatic interactions, while the minimum gelling concentration (MGC) showed only moderate variation, indicating a partial decoupling between early aggregation events and macroscopic gelation behavior. Circular dichroism revealed increasingly cooperative and ordered supramolecular assembly from fluorinated to iodinated peptides, driven primarily by enhanced exciton coupling between aromatic residues. Rheological measurements revealed a nonlinear modulation of hydrogel mechanics across the halogen series, with storage moduli ranging from ∼0.4 kPa for the iodinated system to ∼1.5 kPa for the brominated analogue, reflecting the balance between local supramolecular ordering and higher-order network connectivity. All hydrogels displayed good cytocompatibility toward both NIH/3T3 fibroblasts and HaCaT keratinocytes, maintaining cell viability above 70%. NMR studies of the iodinated analogue provided molecular-level insight into early aggregation, supporting the formation of antiparallel dimeric nuclei as initial assembly intermediates. Overall, these results demonstrate that halogenation provides a tunable and systematic approach to control peptide self-assembly, enabling modulation of supramolecular organization, mechanical properties, and network architecture. This work establishes halogen-dependent aromatic interactions as a key design parameter for the development of peptide hydrogels with tailored properties for potential biomedical and materials applications.