Substrate ectodomain is critical for substrate preference and inhibition of γ-secretase

Understanding the substrate recognition mechanism of γ-secretase is a key step for establishing substrate-specific inhibition of amyloid β-protein (Aβ) production. However, it is widely believed that γ-secretase is a promiscuous protease and that its substrate-specific inhibition is elusive. Here we show that γ-secretase distinguishes the ectodomain length of substrates and preferentially captures and cleaves substrates containing a short ectodomain. We also show that a subset of peptides containing the CDCYCxxxxCxCxSC motif binds to the amino terminus of C99 and inhibits Aβ production in a substrate-specific manner. Interestingly, these peptides suppress β-secretase-dependent cleavage of APP, but not that of sialyltransferase 1. Most importantly, intraperitoneal administration of peptides into mice results in a significant reduction in cerebral Aβ levels. This report provides direct evidence of the substrate preference of γ-secretase and its mechanism. Our results demonstrate that the ectodomain of C99 is a potent target for substrate-specific anti-Aβ therapeutics to combat Alzheimer’s disease. γ-Secretase inhibitors are studied for their potential to treat Alzheimer’s disease, but their use is limited by side effects. Funamoto et al.show that γ-secretase preferentially cleaves substrates with short ectodomains and that inhibitors based on these ectodomains reduce disease-like pathology in mice.