Cloning and Functional Analysis of Skin Host Defense Peptides from Yakushima Tago's Brown Frog () and Development of Serum Endotoxin Detection System.

Amphibian skin is a valuable source of host defense peptides (HDPs). This study aimed to identify HDPs with novel amino acid sequences from the skin of and analyze their functions. cDNAs encoding HDP precursors were cloned and sequenced using RT-PCR and 3'-RACE. The novel HDPs were synthesized to evaluate their antimicrobial activity, antioxidant activity, and cytotoxicity. Antimicrobial activity was evaluated by way of broth microdilution and endotoxin- and β-glucan-binding capacity using an enzyme-linked endotoxin binding assay (ELEBA) and a modified ELEBA, respectively. Nine cDNAs encoding precursors for various HDP families, including temporin, ranatuerin-2, brevinin-1, amurin-9, and a novel yakushimin peptide, were identified. Brevinin-1TYa exhibited antibacterial activity against , and brevinin-1TYa and amurin-9TYa induced morphological changes in and . Yakushimin-TYa, amurin-9TYa, and brevinin-1TYa showed concentration-dependent antibacterial effects against the plant pathogens pv. and subsp. . Amurin-9TYa demonstrated strong binding affinity to lipopolysaccharide, lipoteichoic acid, and β-glucan, exhibited antioxidant activity, and lacked cytotoxicity, making it a promising therapeutic candidate. Moreover, brevinin-1TYa showed strong cytotoxicity, whereas yakushimin-TYa exhibited weak cytotoxicity. These findings highlight the potential of these peptides, particularly amurin-9TYa, for future applications as antimicrobial and therapeutic agents.