Identification and optimisation towards bioactivity study of novel antimicrobial peptides from the skin secretion of Rana dybowskii

Abstract

Antimicrobial peptides (AMPs) are considered as a promising solution to the growing problem of bacterial resistance in clinical medicine. Large amounts of AMPs were identified in the skin secretions of the frog, Rana dybowskii. In the first year, Brevinin-2DYf was characterized alongside its synthetic analogues to explore structure-activity relationships and therapeutic potential. The full-length cDNA precursor of Brevinin-2DYf encodes a 33-residue mature peptide. The peptide 2DYf showed potent activity, particularly analogues 2DYf-2, 2DYf-7, and 2DYf-9, which demonstrated 4-16-fold enhanced efficacy. 2DYf-9 showed higher therapeutic index. Time-killing kinetics confirmed rapid bactericidal effects. Mechanistic studies indicated membrane disruption as the primary mode of action. The antiproliferative studies revealed selective cytotoxicity against H838 cells with minimal toxicity to non-malignant cells.
In year two, Brevinin-1DYc, another peptide derived from Rana dybowskii, demonstrated significant antimicrobial activity, primarily targeting Gram-positive bacteria such as S. aureus and MRSA. Structural predictions and CD spectroscopy indicated that Brevinin-1DYc adopts a dual-helix conformation, with modifications altering hydrophobic interactions. The modified analogues, particularly 2306 and 2307, exhibited improved antibacterial efficacy. Time-killing kinetics showed rapid bactericidal effects, with complete bacterial elimination within minutes at optimal concentrations. Haemolysis and therapeutic index assessments suggested that some modifications reduced toxicity while maintaining antimicrobial potency. Notably, synergism testing revealed that Brevinin-1DYc could enhance the efficacy of antibiotics such as vancomycin and ciprofloxacin against drug-resistant strains.
In year three, Ranatuerin-2YJ and its optimized analogue 2YJc displayed potent, broad-spectrum activity, rapid bactericidal effects, and melittin-like membrane permeabilization. 2YJc also exhibited selective cytotoxicity against H838 lung carcinoma cells with minimal effects on normal keratinocytes.
Collectively, these findings highlight the therapeutic potential of optimized AMPs from Rana dybowskii as dual-function antimicrobial and anticancer agents, with favorable efficacy, selectivity, and synergy profiles.


Thesis embargoed until 31st July 2030.

Date of Award	Jul 2025
Original language	English
Awarding Institution	Queen's University Belfast
Supervisor	Lei Wang (Supervisor), Mei Zhou (Supervisor) & Tianbao Chen (Supervisor)