Tyrosine-capped gold nanoparticles enable cysteine-free peptide loading, enhancing the antipseudomonal efficacy of scorpion-derived AamAP1-Lys-NH2 in a burn wound infection model

Cytotoxicity and instability in complex physiological environments remain key barriers to the clinical translation of antimicrobial peptides (AMPs) especially for infected wounds, which contain protease-rich exudate and are highly sensitive to toxicity that can impede healing. This is despite the strong potential of AMPs to combat antimicrobial resistance, with efficacy against a broad spectrum of pathogens including multidrug-resistant Pseudomonas aeruginosa. Loading AMPs onto gold nanoparticles (AuNPs) offers a promising strategy to protect against degradation, control release, and reduce cytotoxicity, however conventional conjugation methods rely on cysteine residues or linker molecules and typically achieve low peptide loading. Here, we present a robust method for loading the AMP, AamAP1-Lys-NH2, onto tyrosine capped gold nanoparticles (Tyr-AuNPs) that achieves nearly complete peptide loading without the need for cysteine or linker residues, while simultaneously reducing cytotoxicity and conferring resistance to trypsin degradation. Fourier-transform infrared (FTIR) spectroscopy confirmed distinct surface chemistry between Tyr-AuNPs and AMP-coated AuNPs, while confocal microscopy verified peptide adsorption. Cumulative release studies showed slow AMP release in water (20% after 1 hour), and rapid, complete release in PBS (100% after 1 hour), demonstrating tuneable delivery under physiological conditions. In vivo, both AamAP1-Lys-NH2-AuNPs and its all-D-enantiomer significantly improved survival of Galleria mellonella larvae with P. aeruginosa burn wound infections (80-85% survival), compared to 55% with free AMP. Since both enantiomers performed equally well, the additional costs of D-enantiomer synthesis are unwarranted when using AuNP delivery. Overall, this study identifies AamAP1-Lys-NH2-AuNPs as a promising therapeutic candidate for the topical treatment of infected wounds, with the Tyr-AuNP platform enabling efficient peptide loading while preserving antimicrobial activity, enhancing stability and minimizing cytotoxicity.