title: Investigating the effects of positive charge and hydrophobicity on the cell selectivity, mechanism of action and anti-inflammatory activity of a Trp-rich antimicrobial peptide indolicidin.
abstract: To investigate the effects of positive charge and hydrophobicity on the cell selectivity, mechanism of action and anti-inflammatory activity of a Trp-rich antimicrobial peptide indolicidin (IN), a series of IN analogs with Trp-->Lys substitution were synthesized. All IN analogs displayed an approximately 7- to 18-fold higher cell selectivity, compared with IN. IN, IN-1 and IN-2 depolarized (50-90%) the cytoplasmic membrane potential of Staphylococcus aureus close to minimal inhibitory concentration (5-10 microg mL(-1)). However, other IN analogs (IN-3 and IN-4) displayed very low ability in membrane depolarization even at 40 microg mL(-1). Confocal laser-scanning microscopy revealed that IN-3 and IN-4 penetrated the Escherichia coli cell membrane, whereas IN, IN-1 and IN-2 did not enter the cell membrane. In the gel retardation assay, IN-3 and IN-4 bound more strongly to DNA compared with IN, IN-1 and IN-2. These findings suggest that the mechanism of antimicrobial action of IN-3 and IN-4 may be involved in the inhibition of intracellular functions via interference with DNA/RNA synthesis. Unlike IN, all IN analogs did not inhibit nitric oxide production or inducible nitric oxide synthase mRNA expression in lipopolysaccharide-stimulated mouse macrophage RAW264.7 cells, indicating that the hydrophobicity of IN is more important for anti-inflammatory activity in lipopolysaccharide-treated macrophage cells than the positive charge.
authors.affiliation: Department of Bio-Materials, Graduate School, Research Center for Proteineous Materials, Chosun University, Gwangju, Korea.
authors.forename: Yong Hai
authors.initials: YH
authors.lastname: Nan
authors.forename: Ka Hyon
authors.initials: KH
authors.lastname: Park
authors.forename: Yoonkyung
authors.initials: Y
authors.lastname: Park
authors.forename: Young Jin
authors.initials: YJ
authors.lastname: Jeon
authors.forename: Yangmee
authors.initials: Y
authors.lastname: Kim
authors.forename: Il-Seon
authors.initials: IS
authors.lastname: Park
authors.forename: Kyung-Soo
authors.initials: KS
authors.lastname: Hahm
authors.forename: Song Yub
authors.initials: SY
authors.lastname: Shin
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D000890:Anti-Infective Agents / Q000737:chemistry* / Q000494:pharmacology*; D000893:Anti-Inflammatory Agents / Q000737:chemistry* / Q000494:pharmacology*; D023181:Antimicrobial Cationic Peptides / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology*; D002460:Cell Line; D004269:DNA, Bacterial / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D004926:Escherichia coli / Q000187:drug effects; D057927:Hydrophobic and Hydrophilic Interactions; D008264:Macrophages / Q000187:drug effects; D051379:Mice; D016297:Mutagenesis, Site-Directed; D009569:Nitric Oxide / Q000037:antagonists & inhibitors; D011485:Protein Binding; D013211:Staphylococcus aureus / Q000187:drug effects
chemical_list: D000890:Anti-Infective Agents; D000893:Anti-Inflammatory Agents; D023181:Antimicrobial Cationic Peptides; D004269:DNA, Bacterial; C073143:indolicidin; D009569:Nitric Oxide