Synthetic Peptide Antibodies via a Rational Approach Based on Disulfide-Stabilized α-Helical Peptides, for the...

Nuclear Protein 1 (NUPR1) is a ubiquitous protein playing an important role in cancer and acute kidney injury. Its specific targeting by natural or synthetic antibodies like molecularly imprinted polymers (MIPs), is therefore of interest. NUPR1 is an intrinsically disordered protein (IDP), such that it displays a high degree of flexibility and an unstable secondary/tertiary structure, resulting in a continuous fluctuation of its conformation in the free state. These characteristics are not in favor of the creation of homogeneous binding sites during molecular imprinting, so that imprinting using peptide epitopes is investigated. Based on an in silico rational approach, two α-helices from the model structure of NUPR1, as predicted by AlPhaFold, are selected. Two cysteine residues are added at both ends of the epitopes to form a disulfide bond, which provides high stability to the α-helix. The template peptides possess the same 3D structure as the epitope fragments in NUPR1. Consequently, they are effective in producing MIP nanogels that cross-react with high affinity (IC₅₀ 1 nm) only with NUPR1. The work indicates that α-helices, besides the preferred flexible loops, can be considered as viable template epitopes for MIPs, opening new opportunities for the obtention of selective antipeptide MIP nanogels for IDPs.