Lysine scanning identifies analogues of GF-17 with enhanced therapeutic performance.

GF-17, the main antimicrobial domain of LL-37, exhibits optimal amphipathicity and heightened hydrophobicity, leading to increased hemolytic activity. A series of novel GF-17-like antimicrobial peptides (AMPs) were designed using lysine scanning to explore how lysine substitutions at various positions impact the biological activity of GF-17. The results showed that lysine substitutions increased the overall charge of GF-17 while decreasing its overall hydrophobicity. Lysine substitution occur at the hydrophilic surface of the GF-17 α-helix increase the peptide drophobic moment, slightly affect its antimicrobial and hemolytic activity. However, when the lysine substitution occue at the hydrophobic surface of the GF-17, the biological activity of GF-17 changes differs. The results indicate that the position of positively charged lysine residue affects the biophysical properties and selectivity of the peptide. Moreover, GF17-8 was a peptide with high antimicrobial activity and low toxicity in vivo and in vitro. GF17-8 exhibited a potent geometric mean MIC of 7.18 μM and a dramatically improved therapeutic index (27.86), thereby representing a 25-fold enhancement over GF-17, due to lysine substitution on its hydrophobic face. These findings offer valuable insights for designing and optimizing antimicrobial peptides beneficial for developing future antimicrobial agents.