Identification and evaluation of a bioactive peptide, QUB-2032, from the skin secretion of the frog, Odorrana andersonii

Abstract

In recent decades, antibiotic-resistant infections due to the long-term extensive use of antibiotics, have become a major global problem, thus there is an urgent need to develop new drugs. The antimicrobial peptides which are secreted and isolated from the amphibian skin have gained broad attention for they appear to have the potential to replace traditional antibiotics as they are the first defensive line against pathogens. In addition, their unique action mechanism which can target the cell membrane of bacteria and lead to cell destruction may effectively decrease the generation of resistance as found in conventional antibiotic drugs.
This thesis mainly focuses on QUB-2032, which was identified as a member of the Nigrocin-2 family from the skin secretion of a Chinese ranid frog, Odorrana andersonii. The peptide QUB-2032 was isolated from the skin secretion of Odorrana andersonii and identified by the method of ‘shotgun’ cloning and reverse-phase HPLC. Then the peptide was synthesized chemically by means of solid-phase synthesis, purified by HPLC and structurally-confirmed by MALDI-TOF mass spectrometry to obtain sufficient material for the evaluation of multiple biological activities.

The bioactivity assays revealed that QUB-2032 had significant antimicrobial activities against the standard Gram-negative bacterium, Escherichia coli (E. coli ATCC CRM8739) and the standard Gram-positive bacterium, Staphylococcus aureus (S. aureus ATCC CRM6538) which showed a strong MIC and MBC value at a low concentration of only 4 μM. Also the peptide showed weak antimicrobial activity against the yeast Candida albicans (C. albicans ATCC CRM10231) for its MIC value was 64 μM and MBC was 128 μM. In the haemolysis assays, QUB-2032 showed a dose-dependent activity at concentrations from 32 μM to 512 μM, and the degree of haemolysis increased from 9.6% to 97.7%. In the MTT anticancer cell proliferation assay, the peptide also showed significant inhibition against the cancer cell line, NCI-H838, at a concentration of 10-4 M with a value of 98.7%. To conclude, the peptide QUB-2032 showed strong bioactivities against E. coli, S. aureus and NCI-H838 cancer cells and also weak activity against C. albicans. However it showed high cytotoxicity on red blood cells. The peptide could be considered, after optimization by modification, as a potential polypeptide drug for infections in the future.

Thesis is embargoed until 31 December 2028.

Date of Award	Dec 2023
Original language	English
Awarding Institution	Queen's University Belfast
Supervisor	Lei Wang (Supervisor), Tianbao Chen (Supervisor) & Yangyang Jiang (Supervisor)