Antimicrobial peptides from skin secretions of Chinese red belly toad Bombina maxima Ren Lai a , Yong-Tang Zheng b , Ji-Hong Shen a , Guan-Jie Liu b , Hen Liu a , Wen-Hui Lee a , Shao-Zhong Tang b , Yun Zhang a, * a Department of Animal Toxinology, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, People’s Republic of China b Department of Viral Immunology, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China Received 27 July 2001; accepted 23 October 2001 Abstract Two groups of antimicrobial peptides have been isolated from skin secretions of Bombina maxima. Peptides in the first group, named maximins 1, 2, 3, 4 and 5, are structurally related to bombinin-like peptides (BLPs). Unlike BLPs, sequence variations in maximins occurred all through the molecules. In addition to the potent antimicrobial activity, cytotoxicity against tumor cells and spermicidal action of maximins, maximin 3 possessed a significant anti-HIV activity. Maximins 1 and 3 were toxic to mice with LD 50 values of 8.2 and 4.3 mg/kg, respectively. Peptides in the second group, termed maximins H1, H2, H3 and H4, are homologous with bombinin H peptides. cDNA sequences revealed that one maximin peptide plus one maximin H peptide derived from a common larger protein. © 2002 Elsevier Science Inc. All rights reserved. Keywords: Amphibian; Peptide; Antimicrobial; Maximin; Bombina maxima; cDNA; Sequence; HIV 1. Introduction In recent years, it has been widely recognized that many organisms use peptides as part of their host defense systems against microorganisms invading [4,26]. Amphibian skin glands are rich resources of antimicrobial peptides. Up to now, based on their sequence/tridimensional structure char- acteristics, the microbicidal peptides from amphibian skin can be grouped into three broad families [2,5,6,9,19]. The first one contains linear amphipathic helix forming peptides such as magainins and related peptides from African clawed frog Xenopus laevis, bombinin-like peptides (BLPs) from Bombina orientalis and Bombina variegata, and dermasep- tins from South American arboreal frog Phyllomedusa sau- vagei. Another family has four different groups of related peptides isolated from various species of the Ranidae fam- ily, namely brevinins-1 and -2, gaegurins, ranalexin, escu- lentins-1 and -2. Different from the peptides in the first family, they all contain 2 cysteine residues at the C-terminal part that form a disulfide bond. The third one includes temporins that were isolated from Rana temporaria. Tem- porins are composed of only 10 –13 residues and they are the smallest antimicrobial peptides known from amphibian. Recently, another linear, cationic antimicrobial peptide, Kassinatuerin-1, was isolated from the skin of African frog Kassina senegalensis [16]. Kassinatuerin-1 exhibits no se- quence similarity with previous characterized antimicrobial peptides from amphibian skin. In addition to antimicrobial activity, it has been demon- strated that some antimicrobial peptides also possess other potent biological activities. Magainins and their analogues have been shown to have significant antitumor activity [1,7, 20]. In addition, two synthetic magainins A and G were shown to have spermicidal activity. Transmission electron microscopic micrographs showed that both magainins al- tered the plasma membranes of sperm [21]. Moreover, It has been demonstrated that polyphemusins and tachyplesins, two antimicrobial peptides from horse crab, have anti-HIV activity [13]. In this report, we describe the isolation, char- acterization, cDNA cloning and biological activities of nine new antimicrobial peptides from skin secretions of Chinese red belly toad Bombina maxima. Based on their structural similarity, they can be divided into two groups. Five pep- tides in the first group, named maximins 1, 2, 3, 4 and 5, are * Corresponding author. Tel.: +86-871-5194279; fax: +86-871- 5191823. E-mail address: zhangy@mail.kiz.ac.cn (Y. Zhang). Peptides 23 (2002) 427– 435 0196-9781/02/$ – see front matter © 2002 Elsevier Science Inc. All rights reserved. PII: S0196-9781(01)00641-6