Impact of tyrosine substitution on tachyplesin I: Development and characterization of the antimicrobial peptide TP I-Y4

This study aimed to design a novel antimicrobial peptide, TP I-Y4, through the substitution of four cysteines with tyrosines in tachyplesin I (TP I), and to investigate its structure-activity relationship. TP I-Y4 exhibited reduced hydrophobicity and higher β-sheet content than the parent peptide TP I, both in aqueous environments and 50% trifluoroethanol (TFE). TP Ⅰ-Y4 demonstrated a potent, dose-dependent affinity for anionic lipopolysaccharide (LPS), and showed enhanced antimicrobial activities against a broad spectrum of bacteria and fungi with minimal toxicity. Structural modifications in TP I-Y4 increased adsorption to bacterial cell surfaces, resulting in significant reductions in surface electronegativity and increases in surface hydrophobicity than TP Ⅰ. These changes are hypothesized to enable TP Ⅰ-Y4 to inflict more damage to bacterial membranes, as well as liposome membranes, during peptide integration into the phospholipid bilayer. Fluorescence spectroscopy confirmed that both peptides were capable of integrating into the phospholipid bilayer, however, TP Ⅰ-Y4 displayed stronger binding and insertion abilities. These findings indicated that neither disulfide linkage nor Cys is essential for antimicrobial functionality. The reduced hydrophobicity, higher β-sheet content, and more flexibility are proposed as key factors contributing to the superior antimicrobial activity and substantially reduced cytotoxicity of the peptide derivative.