Phosphorylation\nas an Effective Tool to Improve Stability\nand Reduce Toxicity of Antimicrobial Peptides

Developing a straightforward and effective strategy to\nmodify antimicrobial\npeptides (AMPs) is crucial in overcoming the challenges posed by their\ninstability and toxicity. Phosphorylation can reduce toxicity and\nimprove the stability of AMPs. Based on these, we designed a series\nof peptides and their corresponding phosphorylated forms. The results\nshowed that all phosphorylated peptides displayed reduced toxicity\nand enhanced stability compared to their unphosphorylated counterparts.\nAmong them, W<sub>3</sub>BipY<sub>8</sub>-P stood out as the most\npromising peptide, exhibiting similar antibacterial activity as its\nunphosphorylated analog W<sub>3</sub>BipY<sub>8</sub> but with significantly\nreduced hemolytic activity (19-fold decrease), cytotoxicity (3.3-fold\ndecrease), and an extended serum half-life 6.3 times longer than W<sub>3</sub>BipY<sub>8</sub>. W<sub>3</sub>BipY<sub>8</sub>-P exerted\nbactericidal effects by disrupting bacterial membranes. Notably, W<sub>3</sub>BipY<sub>8</sub>-P significantly prolonged the survival of\nbacteria-infected animals while its LD<sub>50</sub> was 4.2 times\nhigher than that of W<sub>3</sub>BipY<sub>8</sub>. These findings\nhighlight phosphorylation as an effective strategy for improving the\nantimicrobial properties of AMPs.