Silyl-Based Alkyne-Modifying Linker for the Preparation of C‑Terminal Acetylene-Derivatized Protected Peptides

A novel linker for the synthesis of C-terminal acetylene-functionalized protected peptides is described. This SAM1 linker is applied in the manual Fmoc-based solid-phase peptide synthesis of Leu-enkephalin and in microwave-assisted automated synthesis of Maculatin 2.1, an antibacterial peptide that contains 18 amino acid residues. For the cleavage, treatment with tetramethylammonium fluoride results in protected acetylene-derivatized peptides. Alternatively, a one-pot cleavage-click procedure affords the protected 1,2,3-triazole conjugate in high yields after purification

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

Analysis of the Mechanism of Action of Potent Antibacterial Hetero-tri-organometallic Compounds: A Structurally New Class of Antibiotics

Two hetero-tri-organometallic compounds with potent activity against Gram-positive bacteria including multi-resistant <i>Staphylococcus aureus</i> (MRSA) were identified. The compounds consist of a peptide nucleic acid backbone with an alkyne side chain, substituted with a cymantrene, a (dipicolyl)­Re­(CO)₃ moiety, and either a ferrocene (FcPNA) or a ruthenocene (RcPNA). Comparative proteomic analysis indicates the bacterial membrane as antibiotic target structure. FcPNA accumulation in the membrane was confirmed by manganese tracing with atomic absorption spectroscopy. Both organometallics disturbed several essential cellular processes taking place at the membrane such as respiration and cell wall biosynthesis, suggesting that the compounds affect membrane architecture. Correlating with enhanced antibacterial activity, oxidative stress was induced only by the ferrocene-substituted compound. The organometallics described here target the cytoplasmic membrane, a clinically proven antibacterial target structure, feature a bactericidal but non-bacteriolytic mode of action and limited cytotoxicity within the limits of solubility. Thus, FcPNA represents a promising lead structure for the development of a new synthetic class of antibiotics