2026 Volume 49 Issue 10 Pages 1686-1689
Kn2-7, an α-helical antimicrobial peptide, enhances the activation of mouse macrophage-like RAW264.7 cells in response to DNA containing unmethylated cytosine-guanine motifs (CpG DNA). We previously showed that high amphipathicity is an important determinant of the CpG DNA response-enhancing activity of short α-helical peptides. However, whether this principle applies to non-α-helical or larger amphipathic molecules is not clear. In the present study, polymyxin B (PMB), a non-α-helical amphipathic antimicrobial peptide, enhanced CpG DNA-induced activation of RAW264.7 cells, whereas PMB nonapeptide, which lacks the hydrophobic tail of PMB, did not. We also synthesized dimeric Kn2-7 peptides, Kn-Gly-Kn, Kn-(Gly)3-Kn, and Kn-(Gly)5-Kn, in which two Kn2-7 molecules are linked by one, three, or five glycine residues, respectively. Kn-(Gly)3-Kn and Kn-(Gly)5-Kn, but not Kn-Gly-Kn, retained CpG DNA response-enhancing activity. Peptide structure prediction using PEP-FOLD3 suggested that, in Kn-Gly-Kn, the hydrophobic faces of the two Kn2-7 domains are oriented toward each other, resulting in a less distinct overall amphipathic molecular organization. In contrast, the longer glycine linkers in Kn-(Gly)3-Kn and Kn-(Gly)5-Kn may increase conformational flexibility, allowing each α-helical Kn2-7 domain to function as an independent amphipathic unit. These findings suggest that amphipathic molecular organization is an important factor in enhancing CpG DNA-induced responses and that such enhancement can also occur with a non-α-helical peptide, as demonstrated for PMB.
