Isolation, characterisation and molecular cloning of a bioactive peptide, QUB 1675, from the defensive skin secretion of Odorrana livida
Amphibians use their unique permeable skin to survive in habitats which are full of microorganisms such as bacteria, fungi and viruses. The frog species of the genus Odorrana harbour a large number of peptides in their skin secretions. In this thesis, a novel peptide containing 14 amino acids (TSRCYVYRLKVVCS) was discovered in the skin secretion of Odorrana livida. It was named systematically as QUB 1675. The sequence of the cDNA encoding the biosynthetic precursor of QUB 1675 was identified by using “shotgun” cloning of a cDNA library generated from the polyadenylated mRNA present in the skin secretion of Odorrana livida. To evaluate the bioactivity of this peptide, QUB 1675 was synthesised by solid phase peptide synthesis (SPPS). The synthetic peptide mixture was purified by reverse phase HPLC and the molecular mass of this peptide was confirmed by MALDI-TOF MS. Antibacterial and antifungal properties of the purified synthetic peptide were evaluated by establishing its MICs on a range of test microorganisms. A haemolysis assay was carried out to test its cytotoxicity. MTT assay was performed on four human cancer cell lines which originated from four different tumour types. The trypsin inhibiting ability and the effects on smooth muscles of the synthetic peptide were also tested.
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