Antimicrobial and Cytolytic Activities and Plausible Mode of Bactericidal Action of the Cell Penetrating Peptide Penetratin and Its Lys‐linked Two‐Stranded Peptide

The cell penetrating peptide, penetratin (RQIKIWFQNRRMKWKK‐NH₂) showed potent antimicrobial activity (MIC: 0.5–4 μm ) without any cytotoxicity against mammalian cells. This study investigated the effect of linking together two peptide chains of penetratin on antimicrobial and cytolytic activities and plausible mode of bactericidal action. Two‐stranded penetratin was prepared by a simultaneous solid‐phase synthesis of the two strands of a single lysine residue attached to the solid support. Two‐stranded penetratin markedly increased cytolytic activity against human erythrocytes and NIH‐3T3 mouse fibroblast cells without a significant effect on antimicrobial activity. This finding suggested that penetratin is active as a monomer to bacterial cells but as an oligomer to mammalian cells. Circular dichroism analysis revealed that the α‐helical content of the membrane‐bound penetratin was unaffected by two‐stranded Lys‐linkage. Penetratin had very weak ability in the depolarization of membrane potential of intact Staphylococcus aureus cells and the fluorescent dye leakage of calcein‐entrapped negatively charged bacterial membrane‐mimicking vesicles. In contrast, two‐stranded penetratin significantly caused membrane depolarization and dye leakage. These results suggest that the two‐stranded penetratin induces a significant change in its mode of bactericidal action from the intracellular‐target mechanism to the membrane‐targeting mechanism.