n-n--4-xylylenebis(pyridinium) and Hemolysis

Plantaricin149a (Pln149a) is a cationic antimicrobial peptide, which was suggested to cause membrane destabilization via the carpet mechanism. The mode of action proposed to this antimicrobial peptide describes the induction of an amphipathic α-helix from Ala7 to Lys20, while the N-terminus residues remain in a coil conformation after binding. To better investigate this assumption, the purpose of this study was to determine the contributions of the Tyr1 in Pln149a in the binding to model membranes to promote its destabilization. The Tyr to Ser substitution increased the dissociation constant (KD) of the antimicrobial peptide from the liposomes (approximately three-fold higher), and decreased the enthalpy of binding to anionic vesicles from -17.2 kcal/mol to -10.2 kcal/mol. The peptide adsorption/incorporation into the negatively charged lipid vesicles was less effective with the Tyr1 substitution and peptide Pln149a perturbed the liposome integrity more than the analog, Pln149S. Taken together, the peptide-lipid interactions that govern the Pln149a antimicrobial activity are found not only in the amphipathic helix, but also in the N-terminus residues, which take part in enthalpic contributions due to the allocation at a lipid-aqueous interface.

Topics: Amino Acid Sequence; Anti-Infective Agents; Bacteriocins; Calorimetry; Circular Dichroism; Hemolysis; Humans; Liposomes; Microbial Sensitivity Tests; Molecular Sequence Data; Naphthalenes; Peptides; Phosphatidylglycerols; Phospholipids; Pyridinium Compounds; Spectrometry, Fluorescence; Structure-Activity Relationship; Tryptophan; Tyrosine

The incorporation of a reduced amide bond, psi(CH(2)NH), into peptide results in an increase in the net positive charge and the perturbation of alpha-helical structure. By using this characteristic of the reduced amide bond, we designed and synthesized novel pseudopeptides containing reduced amide bonds, which had a great selectivity between bacterial and mammalian cells. A structure-activity relationship study on pseudopeptides indicated that the decrease in alpha-helicity and the increase in net positive charge in the backbone, caused by the incorporation of a reduced amide bond into the peptide, both contributed to an improvement in the selectivity between lipid membranes with various surface charges. However, activity results in vitro indicated that a perturbation of alpha-helical structure rather than an increase in net positive charge in the backbone is more important in the selectivity between bacterial and mammalian cells. The present result revealed that the backbone of membrane-active peptides were important not only in maintaining the secondary structure for the interactions with lipid membranes but also in direct interactions with lipid membranes. The present study showed the unique function of a reduced amide bond in cytolytic peptides and a direction for developing novel anti-bacterial agents from cytolytic peptides that act on the lipid membrane of micro-organisms.

Topics: Amides; Amino Acid Sequence; Animals; Anti-Bacterial Agents; Anti-Infective Agents; Bacteria; Candida albicans; Cell Membrane; Chromatography, High Pressure Liquid; Circular Dichroism; Coloring Agents; Erythrocytes; Fluoresceins; Hemolysis; Liposomes; Mice; Microbial Sensitivity Tests; Naphthalenes; Oxidation-Reduction; Peptides; Phospholipids; Protein Structure, Secondary; Pyridinium Compounds; Sodium Dodecyl Sulfate; Static Electricity; Structure-Activity Relationship; Substrate Specificity; Trifluoroethanol