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Title: Peptidomic and bioinformatic studies on bioactive peptides from amphibian skin secretions
Author: Gao, Yitian
ISNI:       0000 0004 6425 3285
Awarding Body: Queen's University Belfast
Current Institution: Queen's University Belfast
Date of Award: 2017
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Phyllomedusa, as a significant genus of South American and Neotropical hylid frogs, has been researched for several years and this has produced abundant bioactive peptides. Most frog species have poisonous skin secretions that can act as a defence against potential predators and pathogens. Therefore, Phyllomedusa frogs have been referred to as a "treasure store” and compared to be the most promising drug sources. Antimicrobial peptides (AMPs), as the most widely isolated group of peptides from skin secretions of Phyllomedusa frogs, have been considered as a novel therapeutic approach to address the serious difficulties in treating antibiotic resistant infections. In addition to their direct antimicrobial activities, AMPs can regulate innate immune responses and can facilitate wound healing and angiogenesis. Phylloseptins are a family of potent AMPs that are widely distributed in the skin secretions of phyllomedusine frogs. The structures of these peptides are relatively conserved, containing 19-21 amino acids with C-terminal amidation, a cationic amphiphilic structure and an a-helical domain. Medusins are a recently discovered family of AMPs in the Phyllomedusinae, with highly-conserved sequences and relatively strong antimicrobial activity against Gram-positive bacteria and fungi, but without obvious cytotoxicity against eukaryotic cells at effective antibacterial concentrations. Dermaseptins are one of the most significant families of amphibian host defence peptides that are usually cationic, contain 10-50 amino acids and inhibit the growth of a wide range of microbes. Here, we report novel peptides from the skin secretion of the Phyllomedusa frogs,Phyllomedusa tarsius and Phyllomedusa sauvagii, identified using a high throughput method combining molecular cloning with mass spectrometry to establish their primary structures. A phylloseptin peptide, a medusin peptide and two dermaseptin peptide-precursor-encoding cDNAs were cloned from defensive skin secretion-derived cDNA libraries by a rapid amplification of cDNA ends technique. R6verse-phase HPLC and tandem mass spectrometry confirmed the presence and primary structure of mature peptide sequences. The secondary structure and physicochemical parameters of each peptide were predicted with using a predictive software system. Cationicity- and amphipathicity- enhanced analogues of the phylloseptin and medusin were by engineered through amino acid substitutions by computational modelling. The synthetic peptides displayed varying degrees of activities. Phylloseptin was active against Staphylococcus aureus and Candida albicans and medusin only against Staphylococcus aureus. However, cationicity-enhanced peptides displayed significant increases in potency and broader spectra of antimicrobial activities. In addition, the appearance of activity against antibiotic-resistant MRSA and biofilms was observed after modification of the structures. Both D-substituted analogues of phylloseptin and medusin showed potent antimicrobial activities as well as stable functional efficiency in the serum. Two dermaseptin peptides exhibited high inhibition against multiple tested microorganisms and a range of cancer cells. These data provide evidence thatAMPs may be candidates as novel antibiotic leads and that targeted modification of a natural AMP template can provide new insights into antibiotic design and development.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available