Functional optimisation and structure-activity relationship studies of helix-loop peptides from the skin secretions of the frogs, Odorrana schmackeri and Amolops wuyiensis

Abstract

Currently, drug-resistant bacterial infections and the increasing incidence of cancer, are global challenges with limited effective therapies. It is therefore urgent to discover and analyse new agents with high efficacy and low side effects. Antimicrobial peptides (AMPs) from amphibian skin secretion play a significant role in the host defence system and have been identified as promising candidates for clinical application against microorganisms and cancer cells. This project aims to isolate and identify novel AMPs from frog skin secretion. Furthermore, to perform a series of modifications to study their structure-activity relationships and to optimise the functions of such helix-loop AMPs.

In Chapter 3, a novel des-Leu2 brevinin family peptide was isolated from the skin secretion of Odorrana schmackeri, and named brevinin-1-OS (B1OS). Several variants were designed to study structure-activity relationships and to improve its bioactivity potency and spectrum. It was found that addition of a leucine residue at position 2, enhanced both antibacterial and anti-cancer activity. Moreover, the incorporation of a D-leucine residue significantly decreased haemolytic activity. These data indicated that N-terminal hydrophobicity and the C-terminal cyclic rana box domain, are critical for the bioactivity of brevinin-1 peptides and propose B1OS-D-L as an attractive therapeutic candidate for clinical development.

In Chapter 4, a novel ranatuerin-2 peptide was isolated and identified from the skin secretion of Amolops wuyiensis and named ranatuerin-2-AW (R2AW). Five analogues were progressively designed to evaluate the role of the rana box and optimise the dual antibacterial and anti-cancer activities of R2AW. The results illustrated that the absence of a disulphide bridge and rana box domain did not affect the antibacterial activity of R2AW. Additionally, a cationicity- and hydrophobicity-enhanced variant, R2AW-LK, which displayed significantly improved antibacterial and anti-cancer activity, was successfully designed. Thus, this study demonstrated that rational design and modification are ideal for optimising the dual activities of ranatuerin-2 peptides and propose R2AW-LK as a promising agent for clinical application.
In Chapter 5, a novel brevinin-2 peptide, brevinin-2-OS (B2OS), was isolated and identified from the skin secretion of Odorrana schmackeri. The modification ideas from Chapters 2 and 4 were merged to design a C-terminal truncated and D-leucine incorporated analogue, t-D-B2OS-NH2, with enhanced antibacterial activity and optimised haemolytic activity. The results suggested that the artificial deletion of the rana box resulted in better antibacterial activity, faster killing efficacy and lower toxicity. Overall, this study indicated that the rana box domain is dispensable in brevinin-2 and ranatuerin-2 helix-loop peptides and proposed the design strategy to optimise activity by incorporating a D-leucine residue on the hydrophobic face.

Thesis embargoed until 31 July 2027

Date of Award	Jul 2022
Original language	English
Awarding Institution	Queen's University Belfast
Supervisor	Mei Zhou (Supervisor) & Lei Wang (Supervisor)