Role of Cationic Side Chains in the Antimicrobial Activity of C18G

Abstract

Antimicrobial peptides (AMPs) have been an area of great interest, due to the high selectivity of these molecules toward bacterial targets over host cells and the limited development of bacterial resistance to these molecules throughout evolution. The peptide C18G has been shown to be a selective, broad spectrum AMP with a net +8 cationic charge from seven lysine residues in the sequence. In this work, the cationic Lys residues were replaced with other natural or non-proteinogenic cationic amino acids: arginine, histidine, ornithine, or diaminopropionic acid. These changes vary in the structure of the amino acid side chain, the identity of the cationic moiety, and the pKa of the cationic group. Using a combination of spectroscopic and microbiological methods, the influence of these cationic groups on membrane binding, secondary structure, and antibacterial activity was investigated. The replacement of Lys with most other cationic residues had, at most, 2-fold effects on minimal inhibitory concentration against a variety of Gram-positive and Gram-negative bacteria. However, the peptide containing His as the cationic group showed dramatically reduced activity. All peptide variants retained the ability to bind lipid vesicles and showed clear preference for binding vesicles that contained anionic lipids. Similarly, all peptides adopted a helical conformation when bound to lipids or membrane mimetics, although the peptide containing diaminopropionic acid exhibited a decreased helicity. The peptides exhibited a wider variety of activity in the permeabilization of bacterial membranes, with peptides containing Lys, Arg, or Orn being the most broadly active. In all, the antibacterial activity of the C18G peptide is generally tolerant to changes in the structure and identity of the cationic amino acids, yielding new possibilities for design and development of AMPs that may be less susceptible to immune and bacterial recognition or in vivo degradation.

Investigation of the structure‐activity relationship in ponericin L1 from Neoponera goeldii

“…The spectral shift is determined by the shift in the spectral barycenter, or center of mass, as a function of titrated lipid vesicles. [ 51 ] The data from these binding experiments are shown in Figure 3, with representative spectra of the free and bound peptides shown in Supplemental Figure S1. All of the peptides exhibited the ability to bind all lipid compositions tested (100% PC; 3:1 PC:PG; 3:1 PE:PG; and 7:3 PC:Cholesterol), but clearly bound with higher affinity to those bilayers containing anionic lipids (note the difference in X‐axis scaling in Panels A and C vs B and D in Figure 3).…”

“…Side chain length and cationic moiety identity have both been shown to dramatically impact characteristics of antimicrobials. [ 51,63‐66 ] Additionally the well‐studied TAT sequence, originally derived from HIV, has shown a necessity for Arg residues to function in membrane translocation. [ 67,68 ] In the L1 background, switching Lys to Arg had minimal effects on MIC and MBC for all strains tested, except P. aeruginosa which had a marked decrease in efficacy.…”

“…This is consistent with previous results on AMPs and peptidomimetic polymers. [ 49,51,64,65 ] However, a recent report by Hodges and coworkers has shown the inclusion of Dap in AMPs can cause a small reduction of antibacterial activity but can have a significant reduction in hemolytic activity, improving the overall therapeutic potential. [ 72 ] The loss of activity from shortening the length of the cationic side chain has been attributed to peptide penetration depth in the bilayer, the Dap side chain forcing a shallower, less disruptive conformation.…”

“…[ 72 ] The loss of activity from shortening the length of the cationic side chain has been attributed to peptide penetration depth in the bilayer, the Dap side chain forcing a shallower, less disruptive conformation. [ 51,64 ] The DQA results indicate that the L1‐X does appear to adopt a shallower orientation in PE:PG membranes (model of bacterial membranes) but shows less of a difference compared to the L1‐K in PC or PC:PG membranes. This is consistent with the best performing sequences against E. coli also exhibiting the greatest OM disruption and lowest Q‐ratio in PE:PG membranes.…”

Antibacterial Activity and Mechanism of Action of Bovine Lactoferricin Derivatives with Symmetrical Amino Acid Sequences

“…For an antimicrobial peptide, its biological activity is closely related to many different structural parameters, including chain length, number of α-helices, hydrophilicity, cationicity, hydrophobicity, and other structural parameters [ 15 ]. In this paper, we mainly consider hydrophobicity and cationicity, due to hydrophobicity and cationicity being directly affected the affinity between AMPs and cell membrane [ 16 , 17 ]. We used the peptide LFcinB18–28 (KCRRWQWRMKK), which has asymmetrical amino acid sequence, as a template; the hydrophobicity amino acids included tryptophan (W), proline (P), phenylalanine (F) and the positively charged amino acids lysine (K) and arginine (R) were used to substitute the original amino acids at the N-terminus and C-terminus, making a peptide with a symmetrical amino acid sequence.…”