Here we describe the isolation of a novel C-terminally amidated octadecapeptide-SVIGCWTKS IPPRPCFVK-amide-that contains a disulphide loop between Cys(5) and Cys(15) that is consistent with a Bowman-Birk type protease inhibitor, from the skin secretion of the Chinese Bamboo odorous frog, Huia versabilis. Named HV-BBI, the peptide is encoded by a single precursor of 62 amino acid residues whose primary structure was deduced from cloned skin cDNA. The precursor exhibits the typical organization of that encoding an amphibian skin peptide with a highly-con served signal peptide, an intervening acidic amino acid residue-rich domain and a single HV-BBI-encoding domain located towards the C-terminus. A synthetic replicate of HV-BBI, with the wild-type K (Lys-8) residue in the presumed P1 position, was found to be a potent inhibitor of trypsin with a K-i just slightly less than 19 nM. Substitution at this site with R (Arg) resulted in a significant reduction in potency (K-i 57 nM), whereas replacement of K with F (Phe) resulted in the complete abolition of trypsin inhibitory activity. Thus, HV-BBI is a potent inhibitor of trypsin and the lysyl (K) residue that occupies the P1 position appears to be optimal for potency of action against this protease. (C) 2008 Elsevier Inc. All rights reserved.