Engineering hybrid lantibiotics yields the highly stable and bacteriocidal peptide cerocin V.

Antimicrobial peptides (AMPs) show promise as alternatives to traditional antibiotics for treating drug-resistant infections. Their adaptability and diverse sequence possibilities allow for rational design by modulating physicochemical determinants to achieve desired biological properties, transforming them into peptides for potential new therapies. Nisin, one of the best-studied AMPs, is believed to have potential to be used as a therapeutic, particularly against antibiotic-resistant bacteria. However, its instability in physiological conditions limits its use in clinical applications and pharmaceutical development. Exploration of new natural variants of nisin has uncovered diverse properties using different domains. Shuffling peptide modules can fine-tune the chemical properties of these molecules, potentially enhancing stability while maintaining or improving antimicrobial activity. In this study, hybrid AMPs were created by combining domains from three unique nisin variants, i.e. nisin A, cesin and rombocin, leading to the identification of a promising variant, named cerocin A, which harbours only 25 amino acids compared to the typical 31-35 amino acid length of nisin. Cerocin A demonstrates potent antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA), approaching that of nisin itself. Cerocin A's mode of action involves a dual mechanism through the combination of two domains, consisting of a small ring/domain (6 amino acids) from the C-terminal end of rombocin attached to the preceding peptide of cesin, changing it from a bacteriostatic to a bactericidal peptide. Further mutation studies identified a new variant, cerocin V, with significantly improved resistance against trypsin degradation, while maintaining high potency. Importantly, cerocin V showed no undesired toxic effects on human red blood cells and remained stable in human plasma. In conclusion, we demonstrate that peptide construction using domain engineering is an effective strategy for manipulating both biological and physicochemical aspects, leading to the creation of novel bioactive molecules with desired properties. These constructs are appealing candidates for further optimization and development as novel antibiotics.