Short Antibacterial Peptides with Significantly Reduced Hemolytic Activity can be Identified by a Systematic l‑to‑d Exchange Scan of their Amino Acid Residues

High systemic toxicity of antimicrobial peptides (AMPs) limits their clinical application to the treatment of topical infections; in parenteral systemic application of AMPs the problem of hemolysis is one of the first to be tackled. We now show that the selectivity of lipidated short synthetic AMPs can be optimized substantially by reducing their hemolytic activity without affecting their activity against methicillin resistant <i>Staphylococcus aureus</i> (MRSA). In order to identify the optimized peptides, two sets of 32 diastereomeric H-^dArg-WRWRW-^lLys­(C­(O)­C_<i>n</i>H_2<i>n</i>+1)-NH₂ (<i>n</i> = 7 or 9) peptides were prepared using a split–split procedure to perform a systematic l-to-d exchange scan on the central WRWRW-fragment. Compared to the all-l C₈-lipidated lead sequence, diastereomeric peptides had very similar antibacterial properties, but were over 30 times less hemolytic. We show that the observed hemolysis and antibacterial activity is affected by both differences in lipophilicity of the different peptides and specific combinations of l- and d-amino acid residues. This study identified several peptides that can be used as tools to precisely unravel the origin of hemolysis and thus help to design even further optimized nontoxic very active short antibacterial peptides

Tuning the Activity of a Short Arg-Trp Antimicrobial Peptide by Lipidation of a C- or N‑Terminal Lysine Side-Chain

The attachment of lipids to <i>C</i>- or <i>N</i>-terminally positioned lysine side-chain amino groups increases the activity of a short synthetic (Arg-Trp)₃ antimicrobial peptide significantly, making these peptides even active against pathogenic Gram-negative bacteria. Thus, a peptide with strong activity against <i>S. aureus</i> (1.1–2 μM) and good activity against <i>A. baumannii</i> and <i>P. aeruginosa</i> (9–18 μM) was identified. The most promising peptide causes 50% hemolysis at 285 μM and shows some selectivity against human cancer cell lines. Interestingly, the increased activity of ferrocenoylated peptides is mostly due to the lipophilicity of the organometallic fragment