Molecular cloning and functional investigation of a novel antimicrobial peptide, QUB-1348, from the skin secretion of the spiny frog, Hylarana spinulosa

Abstract

Antimicrobial peptides (AMPs) are a diverse and potent group of molecules derived from various invertebrate, plant, and animal species. A variety of biological activity, such as antiviral and anticancer properties, make AMPs attractive candidates for the development of novel drugs. The antimicrobial peptide, QUB-1348, which was extracted from the skin secretions of Hylarana spinulosa, is the main topic of this thesis. Based on existing literature and database searches, the sequence and activity of QUB-1348 have not been previously reported, indicating that this peptide is a novel discovery.

Using a cDNA library generated from skin secretions, this study is the first to find the biosynthetic precursor encoding the cDNA of QUB-1348, even though the mature peptide sequence FISGLISGLMKAL-NH2 has been identified before. RP-HPLC was utilised to purify the crude peptide. Using a Tribute peptide synthesiser, QUB-1348 was synthesised and MALDI-TOF mass spectrometry was used to confirm its molecular mass.

Tests for haemolysis, antiproliferation, and antibacterial properties were among the several assays used to evaluate the bioactivities of pure QUB-1348. QUB-1348 showed strong antibacterial action in tests against three model microorganisms: the Gram-positive bacterium, Staphylococcus aureus, the Gram-negative bacterium, Escherichia coli, and the yeast, Candida albicans. The relative MICs were 4 µM, 8 µM, and 32 µM. Furthermore, QUB-1348 has obvious hemolytic activity above the concentration of 32 μm. MTT assay showed that QUB-1348 had anti-proliferation effect on H-838 lung cancer cells, and the average IC50 was 21.36 μM. This indicates that QUB-1348 is still toxic to human cells, but it has obvious antibacterial and anti-proliferative activities. It is hoped that future drug research can apply its biological activity, and reduce its toxicity, so that it can be used in the clinic.

Thesis is embargoed until 31 December 2029.

Date of Award	Dec 2024
Original language	English
Awarding Institution	Queen's University Belfast
Supervisor	Lei Wang (Supervisor), Tianbao Chen (Supervisor) & Mei Zhou (Supervisor)