title: Design of potent 9-mer antimicrobial peptide analogs of protaetiamycine and investigation of mechanism of antimicrobial action.
abstract: Protaetiamycine is an insect defensin, a naturally occurring 43-amino-acid-residue antimicrobial peptide derived from the larvae of the beetle Protaetia brevitarsis. In a previous work that aimed at developing short antibiotic peptides, we designed 9-mer peptide analogs of protaetiamycine. Among them, RLWLAIGRG-NH2 showed good antifungal activity against Candida albicans. In this study, we designed four 9-mer peptide analogs based on the sequence of RLWLAIGRG-NH2, in which Gly or Ile was substituted with Arg, Lys, or Trp to optimize the balance between the hydrophobicity and cationicity of the peptides and to increase bacterial cell selectivity. We measured their toxicity to bacteria and mammalian cells as well as their ability to permeabilize model phospholipid membranes. Substitution of Arg for Gly9 at the C-terminus (9Pbw1) resulted in two- to fourfold improvement in antibacterial activity. Further substitution of Gly7 with Lys (9Pbw2 and 9Pbw4) caused four- to eightfold improvement in the antibacterial activity without increase in cytotoxocity, while substitution of Gly7 with Trp (9Pbw3) increased cytotoxicity as well as antibacterial activity. The peptides 9Pbw2 and 9Pbw4 with the highest bacterial cell selectivity were not effective in depolarizing the membrane of Staphylococcus aureus cytoplasmic membranes and showed almost no leakage of a fluorescent dye entrapped within the vesicles. Gel-retardation experiments indicated that 9Pbw2 and 9Pbw4 inhibited the migration of DNA at concentrations >20 microM. Three positively charged residues at the C-terminus in 9Pbw2 and 9Pbw4 may facilitate effective penetration into the negatively charged phospholipid membrane of bacteria. The results obtained in this study suggest that the bactericidal action of our potent antibacterial peptides, namely 9Pbw2 and 9Pbw4, may be attributed to the inhibition of the functions of intracellular components after penetration of the bacterial cell membrane.
authors: Department of Bioscience and Biotechnology, Konkuk University, Seoul 143-701, Korea.
authors: Soyoung
authors: S
authors: Shin
authors: Jin-Kyoung
authors: JK
authors: Kim
authors: Jee-Young
authors: JY
authors: Lee
authors: Ki-Woong
authors: KW
authors: Jung
authors: Jae-Sam
authors: JS
authors: Hwang
authors: Juneyoung
authors: J
authors: Lee
authors: Dong Gun
authors: DG
authors: Lee
authors: Iksoo
authors: I
authors: Kim
authors: Song Yub
authors: SY
authors: Shin
authors: Yangmee
authors: Y
authors: Kim
mesh_terms: D000595:Amino Acid Sequence; D023181:Antimicrobial Cationic Peptides / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D002942:Circular Dichroism; D004247:DNA / Q000378:metabolism; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D011485:Protein Binding; D013329:Structure-Activity Relationship
chemical_list: D023181:Antimicrobial Cationic Peptides; D004247:DNA