Investigating How Proline Linkers Enhance Hybrid Antimicrobial Peptide Activity

Abstract

To combat increasing antibiotic resistance in bacteria, antimicrobial peptides (AMPs) have been proposed as alternatives to conventional antibiotic treatments. AMPs are naturally found in many organisms, including humans, as part of the innate immune response. One approach to designing novel AMPs is to form a hybrid AMP by connecting two individual AMPs, often with a linker consisting of one or more amino acids. Previous studies have shown that some hybrid AMPs have increased antibacterial activity and reduced cytotoxicity to eukaryotic cells. The amino acid linker and hybrid design used may impact antibacterial activity; however, these effects have not yet been systematically evaluated. This study examines hybrid AMPs formed from two translocating histone-derived antimicrobial peptides (HDAPs): the naturally occurring buforin II (BF2) and the synthesized peptide DesHDAP1. Previous work in our laboratory has shown that proline linkers enhance the activity of these BF2/DesHDAP1 hybrids. To investigate which structural feature of proline contributes to this observed increase in activity, additional BF2/DesHDAP1 hybrids were designed using glycine and N-methyl alanine linkers. Antibacterial activity and circular dichroism spectroscopy results suggest that proline's inability to hydrogen bond contributes more to antibacterial activity than physical rigidity. Molecular dynamics simulations were also performed to compare the hybrids’ structure and interactions with membranes on a molecular level. Ongoing work is investigating the activity of these BF2/DesHDAP1 hybrid AMPs against eukaryotic cells.