Modulating amyloid-β 42 aggregation and neurotoxicity by Kunitz domains and their derived peptides

Alzheimer's disease is associated with the aggregation of amyloid-β42 (Aβ42) into species of varying sizes, with intermediate oligomers being the most neurotoxic. We recently reported that amyloid precursor protein inhibitor (APPI), a Kunitz-type protein, and a cyclic peptide derived from its β-domain reduced Aβ42-mediated neurotoxicity, the former by reducing Aβ42 aggregation and formation of toxic Aβ42 oligomers, and the latter by promoting Aβ42 aggregation to form fibrils rather than the neurotoxic Aβ42 oligomers. To address the question of whether these two inhibition mechanisms are controlled by the structure or the amino acid sequence of the protein/peptide, we exploited three Kunitz-type proteins, bikunin, bovine pancreatic trypsin inhibitor (BPTI) and tissue factor pathway inhibitor (TFPI) - chosen for their similar β-strand-rich structures but different sequences to one another and to APPI - and also short peptides that mimic their β-domains, in either cyclic or linear conformation. In-vitro studies showed that the formation of Aβ42 aggregates was reduced by the three Kunitz-type proteins and by their derived cyclic peptides, but not by the linear counterparts of the cyclic peptides. In SH-SY5Y neuroblastoma cells, the Kunitz-type proteins and the cyclic (but not the linear) peptides reduced the intracellular and extracellular accumulation of Aβ42 aggregates, respectively. Both the Kunitz-type proteins and the cyclic peptides inhibited Aβ42-induced mitochondrial membrane depolarization and reduced Aβ42-mediated apoptosis and cell death. Overall, this study thus reveals the potential of the β-hairpin structure, whether as a segment within the Kunitz-type proteins or isolated as a cyclic peptide, to interact with Aβ42, thereby reducing Aβ42 aggregation and hence its neurotoxicity.