In Silico Selection and Evaluation of Pugnins with Antibacterial and Anticancer Activity Using Skin Transcriptome of Treefrog ().

In order to combat bacterial and cancer resistance, we identified peptides (pugnins) with dual antibacterial l-anticancer activity from the () skin transcriptome through in silico analysis. Pugnins A and B were selected owing to their high similarity to the DS4.3 peptide, which served as a template for their alignment to the transcriptome, as well as their function as part of a voltage-dependent potassium channel protein. The secondary peptide structure stability in aqueous medium was evaluated as well, and after interaction with the () membrane model using molecular dynamics. These pugnins were synthesized via solid-phase synthesis strategy and verified by Reverse phase high-performance liquid chromatography (RP-HPLC) and mass spectrometry. Subsequently, their alpha-helix structure was determined by circular dichroism, after which antibacterial tests were then performed to evaluate their antimicrobial activity. Cytotoxicity tests against cancer cells also showed selectivity of pugnin A toward breast cancer (MFC7) cells, and pugnin B toward prostate cancer (PC3) cells. Alternatively, flow cytometry revealed necrotic cell damage with a major cytotoxic effect on human keratinocytes (HaCaT) control cells. Therefore, the pugnins found in the transcriptome of present dual antibacterial-anticancer activity with reduced selectivity to normal eukaryotic cells.