Self-assembling peptides are being applied both in the biomedical area and as building blocks in nanotechnology. Their applications are closely linked to their modes of self-assembly, which determine the functional nanostructures that they form. This work brings together two structural elements that direct nanoscale self-association in divergent directions: proline as a β-breaker and the β-structure-associated diphenylalanine motif, into a single tripeptide sequence. Amino acid chirality was found to resolve the tension inherent to these conflicting self-assembly instructions. Stereoconfiguration determined the ability of each of the eight possible Pro-Phe-Phe stereoisomers to self-associate into diverse nanostructures, including nanoparticles, nanotapes, or fibrils, which yielded hydrogels with gel-to-sol transition at a physiologically relevant temperature. Three single-crystal structures and all-atom molecular dynamics simulations elucidated the ability of each peptide to establish key interactions to form long-range assemblies (i,e., stacks leading to gelling fibrils), medium-range assemblies (i.e., stacks yielding nanotapes), or short-range assemblies (i.e., dimers or trimers that further associated into nanoparticles). Importantly, diphenylalanine is known to serve as a binding site for pathological amyloids, potentially allowing these heterochiral systems to influence the fibrillization of other biologically relevant peptides. To probe this hypothesis, all eight Pro-Phe-Phe stereoisomers were tested in vitro on the Alzheimer’s disease-associated Aβ(1–42) peptide. Indeed, one nonfibril-forming stereoisomer effectively inhibited Aβ fibrillization through multivalent binding between diphenylalanine motifs. This work thus defined heterochirality as a useful feature to strategically develop future therapeutics to interfere with pathological processes, with the additional value of resistance to protease-mediated degradation and biocompatibility.

The self-assembly process of hydrogen-bonded quinquethiophene–rhodanine derivatives has been explored as a function of solvent and temperature. We demonstrate the divergent supramolecular chirality emerging from a single enantiomer by subtle changes in solvent mixtures and sample preparation protocol. Spectroscopic techniques have proved the presence of aggregates where H-bonding interactions play a crucial role.

Water plays a key role in the irreversible transition from peptide fibrils to plates.

Insulin fibrillation is a problem for diabetic patients that can occur during storage and transport, as well as at the subcutaneous injection site, with loss of bioactivity, inflammation, and various adverse effects. Tripeptides are ideal additives to stabilise insulin formulations, thanks to their low cost of production and inherent cytocompatibility. In this work, we analysed the ability of eight tripeptide stereoisomers to inhibit the fibrillation of human insulin in vitro. The sequences contain proline as β-breaker and Phe–Phe as binding motif for the amyloid-prone aromatic triplet found in insulin. Experimental data based on spectroscopy, fluorescence, microscopy, and calorimetric techniques reveal that one stereoisomer is a more effective inhibitor than the others, and cell live/dead assays confirmed its high cytocompatibility. Importantly, in silico data revealed the key regions of insulin engaged in the interaction with this tripeptide, rationalising the molecular mechanism behind insulin fibril formation reduction.