Orientational and aggregational states of magainin 2 in phospholipid bilayers.

Magainins from Xenopus skin are antimicrobial peptides with broad spectra, and their action mechanisms are considered to be the permeabilization of bacterial membranes. To elucidate their molecular mechanisms, three analog peptides of magainin 2, each having a Trp residue substituted for Phe at the 5th, 12th, or 16th position, were synthesized, and their interactions with acidic phospholipid membranes were investigated by fluorescence. The Trp substitution did not significantly affect the properties of the parent peptide. The binding isotherms of these peptides to the membranes, which were obtained on the basis of fluorescence changes upon membrane binding of the peptides, were sigmoidal, suggesting the association of the bound peptide molecules. A quantitative analysis indicated that the formed aggregate is a dimer. The observation that the initial rate constant of magainin 2 induced leakage of calcein from liposomes was dependent on the fourth power of the peptide concentration demonstrates the formation of a tetrameric pore. A blue shift and intensity enhancement of Trp fluorescence in the presence of the membranes indicate that those Trp residues are buried in the hydrophobic region of the bilayers. Furthermore, the depths of the Trp residues, which were determined using the n-doxylphosphatidylcholine quenching technique, were about 10 A from the bilayer center irrespective of the peptide aggregational state. Thus, it was concluded that the orientation of the magainin 2 alpha-helix is parallel to the membrane surface. A model of the pore formation will be proposed on the basis of these observations.