Preserving antimicrobial efficacy while extending peptide longevity: effects of residue glycosylation

The spread of antibiotic-resistant bacteria has prompted the search for new drugs. Antimicrobial peptides (AMPs) are promising candidates, but their pharmacological application is limited by their poor stability, especially against proteolytic enzymes. A strategy to increase AMPs half-life is the introduction of sugars at key residues, a process termed glycosylation. In this work, the RLK10 peptide was obtained from the GKY10 peptide, by replacing the glutamine residue with an asparagine. Then, it was glycosylated at the asparagine by introducing a N-acetylglucosamine, obtaining the peptide RLK10-NAG. To further increase the stability, the N-terminus was acetylated, producing the Ac-RLK10-NAG peptide. In cellulo assays revealed that RLK10 modifications limitedly affect its antimicrobial activity. However, using POPE/POPG liposomes as a bacterial model membrane, marked differences in their mode of action were found. Stability tests against proteases revealed that the N-acetylglucosamine significantly enhances the stability of the RLK10 sequence. Unexpectedly, the acetylation didn't improve the peptide resistance against proteases, rather it increased its degradation susceptibility. These results demonstrate that glycosylation is an effective strategy to improve the AMPs stability while marginally affecting their biological activity, opening to the possibility of using AMPs in medicine and extending their use into other areas such as food preservation.