Interactions of the novel antimicrobial peptide buforin 2 with lipid bilayers: proline as a translocation promoting factor.
Buforin 2 is an antimicrobial peptide discovered in the stomach tissue of the Asian toad Bufo bufo gargarizans. The 21-residue peptide with +6 net charge shows antimicrobial activity an order of magnitude higher than that of magainin 2, a membrane-permeabilizing antimicrobial peptide from Xenopus laevis [Park, C. B., Kim, M. S., and Kim, S. C. (1996) Biochem. Biophys. Res. Commun. 218, 408-413]. In this study, we investigated the interactions of buforin 2 with phospholipid bilayers in comparison with magainin 2 to obtain insight into the mechanism of action of buforin 2. Equipotent Trp-substituted peptides were used to fluorometrically monitor peptide-lipid interactions. Circular dichroism measurements showed that buforin 2 selectively bound to liposomes composed of acidic phospholipids, assuming a secondary structure similar to that in trifluoroethanol/water, which is an amphipathic helix distorted around Pro(11) with a flexible N-terminal region [Yi, G. S., Park, C. B., Kim, S. C., and Cheong, C. (1996) FEBS Lett. 398, 87-90]. Magainin 2 induced the leakage of a fluorescent dye entrapped within lipid vesicles coupled to lipid flip-flop. These results have been interpreted as the formation of a peptide-lipid supramolecular complex pore [Matsuzaki, K. (1998) Biochim. Biophys. Acta 1376, 391-400]. Buforin 2 exhibited much weaker membrane permeabilization activity despite its higher antimicrobial activity. In contrast, buforin 2 was more efficiently translocated across lipid bilayers than magainin 2. These results suggested that the ultimate target of buforin 2 is not the membrane but intracellular components. Furthermore, buforin 2 induced no lipid flip-flop, indicating that the mechanism of translocation of buforin 2 is different from that of magainin 2. The role of Pro was investigated by use of a P11A derivative of buforin 2. The derivation caused a change to magainin 2-like secondary structure and membrane behavior. Pro(11) was found to be a very important structural factor for the unique properties of buforin 2.
Graduate Schools of Pharmaceutical Sciences and Biostudies, Kyoto University, Japan.
S
S
Kobayashi
K
K
Takeshima
C B
CB
Park
S C
SC
Kim
K
K
Matsuzaki
D000595:Amino Acid Sequence; D000818:Animals; D000900:Anti-Bacterial Agents / Q000737:chemistry* / Q000494:pharmacology*; D023181:Antimicrobial Cationic Peptides; D001665:Binding Sites; D001693:Biological Transport, Active / Q000187:drug effects; D002023:Bufo bufo; D002942:Circular Dichroism; D004058:Diffusion; D004926:Escherichia coli / Q000187:drug effects; D006801:Humans; D066298:In Vitro Techniques; D008051:Lipid Bilayers / Q000737:chemistry*; D054806:Magainins; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000494:pharmacology; D010539:Permeability; D011392:Proline / Q000737:chemistry; D011506:Proteins / Q000737:chemistry* / Q000494:pharmacology*; D029867:Xenopus Proteins
D000900:Anti-Bacterial Agents; D023181:Antimicrobial Cationic Peptides; D008051:Lipid Bilayers; D054806:Magainins; D010455:Peptides; D011506:Proteins; D029867:Xenopus Proteins; C097849:buforin II; C053301:magainin 2 peptide, Xenopus; D011392:Proline