Open AccessRole of Phenylalanine and Valine10 Residues in the Antimicrobial Activity and Cytotoxicity of Piscidin-1
Author(s) -
Eunjung Lee,
Areum Shin,
Ki-Woong Jeong,
Bongwhan Jin,
Hum Nath Jnawali,
Soyoung Shin,
Song Yub Shin,
Yangmee Kim
Publication year - 2014
Publication title -
plos one
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.99
H-Index - 332
ISSN - 1932-6203
DOI - 10.1371/journal.pone.0114453
Subject(s) - antibacterial activity , cytotoxicity , peptide , phenylalanine , biochemistry , amino acid , chemistry , antimicrobial , tryptophan , residue (chemistry) , amphiphile , mtt assay , bacteria , biology , in vitro , genetics , organic chemistry , copolymer , polymer
Piscidin-1 (Pis-1) is a linear antibacterial peptide derived from mast cells of aquacultured hybrid striped bass that comprises 22 amino acids with a phenylalanine-rich amino-terminus. Pis-1 exhibits potent antibacterial activity against pathogens but is not selective for distinguishing between bacterial and mammalian cells. To determine the key residues for its antibacterial activity and those for its cytotoxicity, we investigated the role of each Phe residue near the N-terminus as well as the Val 10 residue located near the boundary of the hydrophobic and hydrophilic sectors of the helical wheel diagram. Fluorescence dye leakage and tryptophan fluorescence experiments were used to study peptide-lipid interactions, showing comparable depths of insertion of substituted peptides in different membranes. Phe 2 was found to be the most deeply inserted phenylalanine in both bacterial- and mammalian-mimic membranes. Each Phe was substituted with Ala or Lys to investigate its functional role. Phe 2 plays key roles in the cytotoxicity as well as the antibacterial activities of Pis-1, and Phe 6 is essential for the antibacterial activities of Pis-1. We also designed and synthesized a piscidin analog, Pis-V10K, in which Lys was substituted for Val 10 , resulting in an elevated amphipathic α-helical structure. Pis-V10K showed similar antibacterial activity (average minimum inhibitory concentration (MIC) = 1.6 µM) to Pis-1 (average MIC = 1.5 µM). However, it exhibited much lower cytotoxicity than Pis-1. Lys 10 -substituted analogs, Pis-F1K/V10K, Pis-F2K/V10K, and Pis-F6K/V10K in which Lys was substituted for Phe retained antibacterial activity toward standard and drug-resistant bacterial strains with novel bacterial cell selectivity. They exert anti-inflammatory activities via inhibition of nitric oxide production, TNF-α secretion, and MIP-1 and MIP-2 production. They may disrupt the binding of LPS to toll-like receptors, eventually suppressing MAPKs-mediated signaling pathways. These peptides may be good candidates for the development of peptide antibiotics with potent antibacterial activity but without cytotoxicity.