Interactions of an antimicrobial peptide, magainin 2, with outer and inner membranes of Gram-negative bacteria

Abstract

Magainin peptides, isolated from Xenopus skin, have broad spectra of antimicrobial activity and low toxicities to normal eukaryotic cells, thus being good candidates for therapeutic agents. The mechanism of action is considered to be the permeabilization of bacterial membranes. A number of studies using lipid vesicles have elucidated its molecular detail. However, their interactions with bacteria are not yet well understood. In this paper, we synthesized several magainin analogs with different charges (0 to +6) and hydrophobicities, and systematically studied their interactions with the outer and inner membranes of three species of Gram-negative bacteria (Escherichia coli, Acinetobacter calcoaceticus, Proteus vulgaris). The treatment of the E. coli cells with native magainin 2 (+4) immediately induced the efflux of the intracellular K+ ions and the cell death. A number of blebs were formed on the bacterial surface and the outer membrane became leaky. An increase in the peptide's positive charge enhanced the outer membrane permeabilization and the bactericidal activity. The cationic peptides also effectively permeabilized the inner membranes rich in acidic phospholipids, indicating the importance of electrostatic interactions. Substitution of Trp for Phe simultaneously increased the bactericidal activity and the hemolytic activity. A strategy to develop potent antimicrobial peptides was discussed on the basis of these results.

Translocation of Analogues of the Antimicrobial Peptides Magainin and Buforin across Human Cell Membranes

“…Addition of the peptide to bacterial cells immediately induces the permeabilization of both outer and inner membranes, leading to cell death (9,10). A fluorescent magainin peptide mainly associates with bacterial membranes and does not enter cells (10), although native magainin 2 was recently reported to enter cells (11).…”

“…The information on cell penetration available is limited to dermaseptins (13)(14)(15), PR-39 (16), and protegrin 1 (17), although many investigators examined hemolytic activity (9,18) and cytotoxicity (19 -22). In contrast, short Arg-rich cationic peptides have recently been shown to enter mammalian cells by an energy (ATP)-independent, non-endocytotic pathway and have been utilized as vectors for delivering membrane-impermeable drugs, oligonucleotides, and proteins into cells (23).…”

Antimicrobial peptides from amphibians

“…For example, brevinin-1E from R. esculenta has a HC 50 value (the concentration producing 50% hemolysis) -1 mM and the HC 50 value of magainin 2 is as high as 1000 mM (1,93). As a result, discovery of less toxic antimicrobial peptides either from natural resources or designed from current AMP databases is somewhat necessary for the pharmaceutical application of AMPs.…”

“…When the peptide/lipid ratio exceeds a certain threshold value, the AMPs in the bacterial membrane will form pores. Three models including the barrel-stave model (107,109,110), carpet-like model (111) and toroidal pore model (93,107) are proposed for the pore-forming mechanism.…”

“…The inner diameter is approximately 3.0-5.0 nm and the external diameter is approximately 7.0-8.4 nm. Each channel is formed by approximately 4-7 magainins and 90 lipid molecules (122)(123)(124). iv.…”