title: Functional peptidomics of amphibian skin secretion: A novel Kunitz-type chymotrypsin inhibitor from the African hyperoliid frog, Kassina senegalensis.
abstract: Amphibian skin secretions are, for the most part, complex peptidomes. While many peptide components have been biologically- and structurally-characterised into discrete "families", some of which are analogues of endogenous vertebrate regulatory peptides, a substantial number are of unique structure and unknown function. Among the components of these secretory peptidomes is an array of protease inhibitors. Inhibitors of trypsin are of widespread occurrence in different taxa and are representative of many established structural classes, including Kunitz, Kazal and Bowman-Birk. However, few protease inhibitors with activity against other specific proteases have been described from this source. Here we report for the first time, the isolation and structural characterisation of an inhibitor of chymotrypsin of Kunitz-type from the skin secretion of the African hyperoliid frog, Kassina senegalensis. To this end, we employed a functional peptidomic approach. This scheme involves fractionation of the peptidome, functional end-point screening, structural characterisation of resultant actives followed by molecular cloning of biosynthetic precursor-encoding cDNA(s). The novel mature and active polypeptide identified consisted of 62 amino acid residues (average molecular mass 6776.24 Da), of which 6 were positionally-conserved cysteines. The P(1) position within the active site was occupied by a phenylalanyl residue. Bioinformatic analysis of the sequence using BLAST, revealed a structural similarity to Kunitz-type chymotrypsin inhibitors from other organisms, ranging from silkworms to snakes.
authors.affiliation: Natural Drug Discovery group, School of Pharmacy, Queen's University, Belfast BT9 7BL, Northern Ireland, UK.
authors.forename: Hui
authors.initials: H
authors.lastname: Wang
authors.forename: Lei
authors.initials: L
authors.lastname: Wang
authors.forename: Mei
authors.initials: M
authors.lastname: Zhou
authors.forename: Mu
authors.initials: M
authors.lastname: Yang
authors.forename: Chengbang
authors.initials: C
authors.lastname: Ma
authors.forename: Tianbao
authors.initials: T
authors.lastname: Chen
authors.forename: Yingqi
authors.initials: Y
authors.lastname: Zhang
authors.forename: Martin
authors.initials: M
authors.lastname: Zeller
authors.forename: Martin
authors.initials: M
authors.lastname: Hornshaw
authors.forename: Chris
authors.initials: C
authors.lastname: Shaw
mesh_terms: D000663:Amphibians; D000818:Animals; D001001:Anura; D002918:Chymotrypsin / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry / Q000494:pharmacology*; D011480:Protease Inhibitors / Q000737:chemistry / Q000494:pharmacology; D012867:Skin / Q000737:chemistry*
chemical_list: D010455:Peptides; D011480:Protease Inhibitors; D002918:Chymotrypsin