Metal-mediated nanobody assemblies as potent alleviators of human islet amyloid polypeptide aggregation

Abstract

The misfolding and aggregation of peptides and proteins into β-sheet-enriched amyloid fibrils has been implicated in many human diseases. Inhibition of protein aggregation by engineered nanobodies has shown great promise in the treatment of amyloid-associated diseases. Taking type 2 diabetes associated human islet amyloid polypeptide (IAPP) aggregation as a model system, we generated a nanobody inhibitor by grafting the IAPP peptide fragment into the complementary determining region of a parent nanobody to inhibit IAPP aggregation through homotypic interactions. In addition, we developed a facile fabrication strategy to amplify the inhibitory effects of the designed nanobody inhibitor on IAPP aggregation. By coordinating a metal cation Zn²⁺ with a histidine-tag-fused nanobody inhibitor M1, the achieved nanobody assemblies M1@Zn²⁺ can significantly enhance the binding affinity between IAPP and M1 through the multivalent effects. At low substoichiometric concentrations (20 : 1 IAPP : nanobody molar ratio), M1@Zn²⁺ are capable of efficiently inhibiting IAPP aggregation, alleviating IAPP-induced cytotoxicity and downregulating ROS generation. This strategy represents an innovative attempt to design high-efficiency amyloid antibody inhibitors with enhanced therapeutic effects for the treatment of amyloid diseases.