Ultrashort self-assembling peptides as antimicrobial agents: structure-function relationships and applications

<p>Antimicrobial resistance was described by the World Health Organization as one of the main public health concerns of the 21st century that threatens the effective prevention and treatment of an increasing range of common infections. These data led the scientific community to engage in research on alternative strategies to traditional small-molecule antibiotics. Antimicrobial peptides are produced by various organisms as part of their normal immune response. A growing number of these peptides have been shown to self-assemble into nanostructures that are thought to play a role in their antimicrobial activity and can be exploited to create biomaterials. While most studies to date have focused on long peptide sequences and proteins, recent works demonstrated that short peptide sequences also selfassemble and display antimicrobial activity. For instance, battacin derivatives composed of only 3-5 amino acid residues can self-assemble, interact with bacterial membranes, and display potent antimicrobial activity.</p> <p>This PhD thesis focuses on the rational design and microbiological/biophysical characterization of ultrashort antimicrobial peptides that present potential as antimicrobial biomaterials. Chapters 1 and 2 entail a comprehensive review on the key topics of the research, followed by a description of the techniques and methods used in the project. Chapters 3, 4 and 5 reports the different approaches used in the design of peptides, followed by their antimicrobial and self-assembling characterization, while Chapter 6 describes the characterization of the interaction with model membranes, hemolytic activity and enzymatic stability of the active designed peptides.</p> <p>Approximately 20 peptides were designed and allocated in 3 families: 1) Indolicidin (ditryptophan); 2) Substance P (diphenylalanine); and 3) Neuropeptide Y (dityrosine). Combining rational design with microbiological characterization, a potent peptide named Priscilicidin (Fmoc-WWRR-NH2) was created. Important correlations between activity and peptide antimicrobial motifs, including, net charge, hydrophobicity, polarity and amphiphilicity was demonstrated.</p> <p>Using a set of techniques we verified that some of these designed peptides, to mention Priscilicidin, self-assembled into liquid crystalline beta-sheet fibrillar nanostructures in 2 aqueous media, resulting in the formation of hydrogels. Other peptides self-assembled into stiff nanotubes/nanofibers.</p> <p>Investigation of the peptide interaction with bacterial, mammalian and fungal model membranes combined with hemolytic activity assay and calculated therapeutic index revealed that our cationic peptides interacted differently with mammalian and bacterial/fungal membranes. The lead peptide Priscilicidin, presented a lower hemolytic capacity than the control indolicidin while preserving the antimicrobial activity, hence with a higher selectivity towards microbial cells.</p> <p>Promising applications for the obtained antimicrobial hydrogels include topical formulations, while stiff nanotubes can be potentially used on the synthesis on antimicrobial topologies.</p>