Discovery and characterization of a high-affinity G-quadruplex binding peptide via mRNA display

G-quadruplexes (G4s) are non-canonical secondary structures of nucleic acids that play crucial roles in gene expression, and their dysregulation has been implicated in various diseases. Therefore, development of G4-binding molecules, including peptides and proteins, is required to modulate G4-dependent biological processes for therapeutic purposes. In this study, a novel G4-binding peptide with high affinity for the G4 structure was developed using directed evolution based on mRNA display. The identified peptide, LP7, exhibited a preferential affinity for parallel G4 structures. Dimerization of LP7 significantly enhanced its binding to hTERC rG4 by approximately 70-fold, with a K_d of 7 nM. Analysis of the sequence-activity relationship revealed that both the basic and aromatic amino acid residues of the peptide are critical for its binding affinity to G4. Functional assays confirmed that LP7 inhibits reverse transcription in a G4-dependent manner by binding to the rG4 region. This study demonstrates the successful application of the mRNA display platform for discovering novel G4-binding peptides. A detailed characterization of LP7 provides valuable insights into the molecular interactions that govern G4 recognition. These findings highlight the potential of G4-binding peptides as tools for targeting and regulating G4-mediated gene functions, offering a promising avenue for the development of G4-dependent therapeutic strategies in the future.