Effects of Hydrophobic Peptoid Substitutions on the Bacterial Cell Selectivity and Antimicrobial Activity of Piscidin 1

Abstract

Piscidin 1 (Pis-1) from hybrid striped bass possesses antibacterial activities with high cytotoxicity. Previously, we reported that Pis-1[NkG], where Gly 8 was substituted with positively charged Lys peptoid in Pis-1, shows high antibacterial activity with more potent bacterial cell selectivity than parent Pis-1 and Pis-1[PG] with Pro substitution at Gly 8 . Here, we investigated the effects of hydrophobic peptoid substitutions on antimicrobial activity and cytotoxicity of Pis-1. We designed Pis-1[NaG] and Pis-1[NlG] peptides, where Gly 8 was replaced with alanine or leucine peptoids. Although hydrophobicity was increased in Pis-1[NaG] and Pis-1[NlG] compared with that in Pis-1[NkG], Pis-1, and Pis-1[PG], these peptides retained high antibacterial activity and showed no hemolytic activity when used at 100 μM. All peptoid-substituted peptides penetrated well the bacterial membrane and localized inside of the cytoplasm, while Pis-1 and Pis-1[PG] depolarized membrane severely. The flexible hinge structure formed by peptoid residues compared with the rigid bent structure formed by proline may explain their differential mechanisms of action. Most hydrophobic peptide, Pis-1 [NlG], showed the most efficient inhibition of LPS (lipopolysaccharide)-induced nitric oxide production, indicating that it can be a potent antimicrobial peptide with anti-inflammatory activities. Peptoid substitution may facilitate the development of potent peptide drugs with bacterial cell selectivity.

Antibacterial Activity of AI-Hemocidin 2, a Novel N-Terminal Peptide of Hemoglobin Purified from Arca inflata

“…Piscidin 1, a peptide purified from hybrid striped bass, possesses significant antibacterial action as well as cytotoxic activity [ 45 ]. The mechanism of action of its bactericidal and fungicidal effects was operated via targeting and disrupting the bacterial and fungal membranes [ 46 , 47 ]. However, the application of piscidin 1 has been limited on account of its cytotoxic effects.…”

Advances in Development of Antimicrobial Peptidomimetics as Potential Drugs

“…Likewise, replacement of Pro residues in Trp/Pro-rich AMPs appears to constitute a promising approach for development of cell-selective AMPs with intracellular mechanisms [ 196 ]. In most cases the modification with α-peptoid residues confers broad-spectrum antibacterial activity against common Gram-negative and Gram-positive pathogens [ 191 , 192 , 193 , 194 , 195 , 196 , 197 , 198 ]. With few exceptions preparation involves solid-phase peptide synthesis (SPPS) where Fmoc-protected peptoid building blocks are used at the appropriate positions.…”

“…Piscidin 1 from a hybrid of striped bass is an antibacterial and cytotoxic α-helical peptide (22 residues) for which analogs with peptoid residues (FFHHIFRXIVHVGKTIHRLVTG, X = P, N Ala, N Leu or N Lys) were designed [ 193 , 197 ]. The hydrophobicity of the N Ala- and N Leu-substituted analogs was higher than that of the N Lys-containing analog, but all peptides retained high antibacterial activity (MICs of 1–8 µM) against a panel of Gram-negative and Gram-positive bacteria, and were devoid of hemolytic activity at a concentration of 100 μM, whereas the original AMP was hemolytic at 3 µM.…”

“…While Pis-1[PG] had a rigid bent structure at the central Pro, Pis-1[NkG] was found to exist as two conformers with a flexible hinge structure at N Lys [ 193 ]. Confocal laser-scanning microscopy revealed that all fluorophore-labelled peptoid-containing analogs penetrated the bacterial membrane and accumulated in the cytoplasm, whereas Pis-1[PG] localized to the surface and displayed extensive membrane depolarization of S. aureus [ 193 , 197 ]. The different conformations induced by Pro and Gly replacement by a peptoid residue may account for the distinct killing mechanisms of the resulting analogs.…”

“…The different conformations induced by Pro and Gly replacement by a peptoid residue may account for the distinct killing mechanisms of the resulting analogs. Thus, introduction of an N Lys residue provided simultaneous conformational flexibility and increased cationicity in the hinge region, which facilitated penetration of the bacterial cell membrane and conferred the highest bacterial cell selectivity to Pis-1[NkG] [ 193 , 197 ]. Moreover, the most hydrophobic N Leu-substituted analog exhibited inhibition of LPS-induced nitric oxide production at a concentration of 2 µM [ 197 ].…”