Healthcare-associated infections resulting from bacterial attachment and biofilm formation on medical implants are posing significant challenges in particular with the emergence of bacterial resistance to antibiotics. Here, we report the design, synthesis and characterization of self-assembled nanostructures, which integrate on their surface antibacterial peptides. The antibacterial WMR peptide, which is a modification of the native sequence of the myxinidin, a marine peptide isolated from the epidermal mucus of hagfish, was used considering its enhanced activity against Gram-negative bacteria. WMR was linked to a peptide segment of aliphatic residues (AAAAAAA) containing a lipidic tail (C19H38O2) attached to the ε-amino of a terminal lysine to generate a peptide amphiphile (WMR PA). The self-assembly of the WMR PA alone, or combined with coassembling shorter PAs, was studied using spectroscopy and microscopy techniques. The designed PAs were shown to self-assemble into stable nanofiber structures and these nanoassemblies significantly inhibit biofilm formation and eradicate the already formed biofilms of Pseudomonas aeruginosa (Gram-negative bacteria) and Candida albicans (pathogenic fungus) when compared to the native WMR peptide. Our results provide insights into the design of peptide based supramolecular assemblies with antibacterial activity, and establish an innovative strategy to develop self-assembled antimicrobial materials for biomedical applications.