Heterochirality and Halogenation Control Phe-Phe Hierarchical Assembly

Diphenylalanine is an amyloidogenic building block that can form a versatile array of supramolecular materials. Its shortcomings, however, include the uncontrolled hierarchical assembly into microtubes of heterogeneous size distribution and well-known cytotoxicity. This study rationalized heterochirality as a successful strategy to address both of these pitfalls and it provided an unprotected heterochiral dipeptide that self-organized into a homogeneous and optically clear hydrogel with excellent ability to sustain fibroblast cell proliferation and viability. Substitution of one l-amino acid with its d-enantiomer preserved the ability of the dipeptide to self-organize into nanotubes, as shown by single-crystal XRD analysis, whereby the pattern of electrostatic and hydrogen bonding interactions of the backbone was unaltered. The effect of heterochirality was manifested in subtle changes in the positioning of the aromatic side chains, which resulted in weaker intermolecular interactions between nanotubes. As a result, d-Phe-l-Phe self-organized into homogeneous nanofibrils with a diameter of 4 nm, corresponding to two layers of peptides around a water channel, and yielded a transparent hydrogel. In contrast with homochiral Phe-Phe stereoisomer, it formed stable hydrogels thermoreversibly. d-Phe-l-Phe displayed no amyloid toxicity in cell cultures with fibroblast cells proliferating in high numbers and viability on this biomaterial, marking it as a preferred substrate over tissue-culture plastic. Halogenation also enabled the tailoring of d-Phe-l-Phe self-organization. Fluorination allowed analogous supramolecular packing as confirmed by XRD, thus nanotube formation, and gave intermediate levels of bundling. In contrast, iodination was the most effective strategy to augment the stability of the resulting hydrogel, although at the expense of optical transparency and biocompatibility. Interestingly, iodine presence hindered the supramolecular packing into nanotubes, resulting instead into amphipathic layers of stacked peptides without the occurrence of halogen bonding. By unravelling fine details to control these materials at the meso- and macro-scale, this study significantly advanced our understanding of these systems.