Inhibitory effect of short cationic homopeptides against Gram-positive bacteria

In the selection or design of antimicrobial peptides, the key role played by cationic amino acids and chain length on the inhibitory potency and specificity is not clear. A fundamental study was conducted using chemically synthesized homopeptides of L-Lys and L-Arg ranging from 7 to 14 residues. Their effect on growth inhibition was evaluated over a wide range of Gram-positive bacteria at different levels of concentration. Interestingly, at lower concentrations (10 μM), Lys homopeptides with odd number of residues, especially with 11 residues, showed a broader inhibitory activity than those with even number of residues. At higher peptide concentrations (>20 μM), the inhibitory activity of Lys homopeptides was directly related to the number of residues in the chain. In contrast, Arg homopeptides, at lower concentrations, did not exhibit a defined pattern of bacterial inhibition related to the number of residues; however, at higher concentrations (>20 μM), the inhibitory effects were more pronounced. Lys homopeptides at concentrations up to 300 μM showed a remarkably lower toxicity against CHSE-214 cells. Arg homopeptides exhibited negligible cytotoxicity up to chain length of 11 residues at concentrations lower than 100 μM, but an abrupt increase in toxicity resulted when the peptide chain length reached 12 amino acid residues and higher concentrations. All synthesized homopeptides displayed characteristic polyproline II helix conformation in both buffer and liposomes, as shown by CD spectroscopy. This result suggests that short Lys homopeptides with an odd number of residues (9 and 11) have a broad spectrum of activity against Gram-positive bacterial cells compared with Arg homopeptides, which in turn showed a considerably higher selectivity toward those cells. By investigating the differences between Lys and Arg homopeptides, this study contributes to the understanding of their mechanism of growth inhibition and selectivity. Thus, it provides further guidelines for a rational design of short antimicrobial peptides. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd

Arginine Homopeptide of 11 Residues as a Model of Cell-Penetrating Peptides in the Interaction with Bacterial Membranes

Cell-penetrating peptides rich in arginine are good candidates to be considered as antibacterial compounds, since peptides have a lower chance of generating resistance than commonly used antibiotics. Model homopeptides are a useful tool in the study of activity and its correlation with a secondary structure, constituting an initial step in the construction of functional heteropeptides. In this report, the 11-residue arginine homopeptide (R11) was used to determine its antimicrobial activity against Staphylococcus aureus and Escherichia coli and the effect on the secondary structure, caused by the substitution of the arginine residue by the amino acids Ala, Pro, Leu and Trp, using the scanning technique. As a result, most of the substitutions improved the antibacterial activity, and nine peptides were significantly more active than R11 against the two tested bacteria. The cell-penetrating characteristic of the peptides was verified by SYTOX green assay, with no disruption to the bacterial membranes. Regarding the secondary structure in four different media—PBS, TFE, E. coli membrane extracts and DMPG vesicles—the polyproline II structure, the one of the parent R11, was not altered by unique substitutions, although the secondary structure of the peptides was best defined in E. coli membrane extract. This work aimed to shed light on the behavior of the interaction model of penetrating peptides and bacterial membranes to enhance the development of functional heteropeptides

Proline and alanine scanning analysis and structure-function relationships of the 11-residue lysine homopeptides

In a previous work we demonstrated that Lys homopeptides of odd number of residues were able to inhibit a wide range of both Gram (+and Gram-negative bacteria, especially with 11-residues, as opposite toLys homopeptides with an even number of residues [1].Here, we used alanine and proline scanning analogs to elucidate the contribution of the side chain of each ε–aminogroup to the antibacterial and cytotoxic activity of the peptides. Hence, 11 peptides of scan Ala and the same number of scan Pro were synthesized by Fmoc solid-phase synthesis and subsequently tested for antibacterial activity against a wide range of bacteria. Lys residues at either the C-or N-terminal end of the peptide played a prominent role for bacterial inhibition compared to Lys residues located at the center of the chain, as demonstrated by Ala exchange. In general, substitution of Lys for Pro did not substantially change the bacterial inhibition pattern. Ala substitutions at any position of the homopeptide showed amore significant change of the polyproline II (PPII) structure compared to Pro substitutions. Indeed, Pro substitutions showed an increased propensity to form PPII structures. Besides providing insight into the biophysical attributes and the critical positions within the homopeptide sequence, which govern the antibacterial activity of cationic homopeptides, our studies assist in recognizing the key roles played by different amino acids in the antibacterial activity