Peptide Nucleic Acids (PNAs) in STD NMR experiments: a simple and valuable tool for studying HuR-small molecule complexes

Ribonucleic acid (RNA)-binding proteins (RBPs) play a key role in regulating RNA stability, fate, function, gene expression, post-transcriptional modifications, and cellular activities. Among the various RBPs identified to date, the Hu proteins have been the most extensively studied. Specifically, HuR influences several cellular processes, including cell proliferation, differentiation, and stress response, and it is frequently overexpressed in various solid tumors. Several HuR ligands have been identified so far, highlighting the druggability of such a protein. To discover the novel HuR–RNA interfering agents, biophysical assays represent a promising approach. To overcome limitations for RNA manipulation, in this work, we explored the use of PNA (peptide nucleic acid) as an RNA analogue in interaction studies. Molecular modeling simulation revealed the ability of aegPNA to bind HuR and, therefore, the synthesis of the designed PNA was conducted. The saturation transfer difference (STD) nuclear magnetic resonance (NMR) technique was adopted to evaluate the ability of HuR ligands to interfere with the HuR–PNA complex, comparing the obtained results with RNAs. Our results evidenced that PNA may be considered a simple and valuable tool to analyze the interaction and interfering properties of HuR ligands by STD-NMR, thus improving the precision and reliability of the approach.