Purification, Characterization, CDNA Cloning And Function Analysis Of Antimicrobial Peptides From Skin Tissue Of Amolops Chunganensis

To date, there are45species of Amolops in the world, and19of them are widelydistributed in tropical and subtropical China. The Chungan torrent frog, Amolopschunganensis is medium sized frog inhabiting woodland areas of mountains in western andsouthern China with large population size. This species is primarily nocturnal and prevalenton the fringes of lakes, ponds, and slow moving rivers between altitudes of700m and1800mduring the reproductive season.This study has purified and characterized four novel antimicrobial peptides from the skinsecretions of Chungan torrent frog, Amolops chunganensis, Sequence analysis indicates thatthey belong to temporin, esculentin-2, and palustrin-2families. Therefore, these peptides werenamed as temporin-CG1(FLPFVGNLLKGLL-NH2), temporin-CG2(FFPIVGKLLSGLF-NH2), esculentin-2CG1(SLFSIFKTAAKFVGKNLLKQAGKAGLETLACKAKNEC), and palustrin-2CG1(GLWNTIKEAGKKFAINVLDKIRCGIAGGCKT) respectively.cDNAs encoding the fivenovel antimicrobial peptide precursors have also been cloned through screening the skincDNA library of A. chunganensis. The actual sequences and characteristics of mature AMPswere analyzed by RP-HPLC and LC-MS/MS-based proteomics approaches in combination offour different protein digestion processes and by LTQ XL.While all the peptides displaypotent and broad-spectrum antimicrobial activities against both Gram-positive andGram-negative bacteria, they show weak hemolytic activity against mammalian red bloodcells in vitro. Therefore, these novel peptides were promising candidates for use asantimicrobial drugs in future.