The development of novel antimicrobials is a top priority to address the growing epidemic of multidrug-resistant pathogens. Since cationic nonamphiphilic star-shaped antimicrobials are promising molecular scaffolds that provide a high charge density in binding anionic bacterial bilayers, this research aimed to further increase their membrane perturbation capability by introducing guanidinium groups to the antimicrobials via enhancing membrane insertion. In particular, computational simulation and experimental investigations revealed that our designed guanidinium-rich alternating copolypeptide, four-armed poly(arginine-alt-glycine), can interact with both the headgroups and unsaturated tails of phospholipids in bacterial membranes through multiple interactions, including electrostatic, cation-π, and T-shaped π-π interactions, allowing it to penetrate deeper inside the biologically inaccessible high-energy barrier of the hydrophobic lipid bilayer interior to cause membrane permeabilization and precipitation of the bacterial cytoplasm. Furthermore, glycine was observed to have a unique effect in enhancing the performance of arginine-based copolypeptide. Four-armed poly(arginine-alt-glycine) exhibited broad-spectrum antimicrobial activity, high bactericidal efficiency, and negligible hemolysis. The in vivo antibacterial performance of the copolypeptide was superior to that of doxycycline in a mouse model of Pseudomonas aeruginosa skin infection, accompanied by negligible local and systemic toxicity. Our results demonstrate that this guanidinium-rich, nonamphiphilic, star-shaped structure may promote the development of next-generation antimicrobials.
Keywords: antimicrobial agent; guanidinium-rich copolypeptides; membrane perturbation and permeability; nonamphiphilic star-shaped structure; peptide−membrane interactions.