Rational modification and bioactivity evaluation of a bioactive peptide, QUB-2607, from the skin secretion of Rana pipiens

  • Yao Li

Student thesis: Doctoral ThesisDoctor of Philosophy


The aim of this research was to identify a novel bioactive peptide from the skin secretion of Rana pipiens, evaluate the bioactivities of the its synthetic replicate and explore the action mechanism following the identification of its full length of the prepropeptide-encoding cDNA by molecular cloning. Afterwards, novel peptide QUB-2607 was identified and then a series of modified peptides were designed to improve its bioactivity. Furthermore, the relevance between the Rana box motif of the brevinin-1 subfamily peptides and their bioactivity were also explored.

With the worldwide emergency of super-resistant bacteria, antimicrobial peptides (AMPs) have been considered antibiotic candidates because of their broad-spectrum antimicrobial activity, low risk of causing drug-resistance and strong antibacterial activity. Furthermore, skin and soft tissue infections (SSTIs) have been growing continuously, and Methicillin-resistant Staphylococcus aureus-infected (MRSA) SSTIs occupied over half of the cases.

In Chapter 3, a novel peptide, QUB-2607, was isolated from the skin secretion of Rana pipiens. The sequence of QUB-2607 was translated by molecular cloning. After alignment of QUB-2607 with other peptides recorded in the antimicrobial peptide database (APD), QUB-2607 was identified as belonging to the brevinin-1 family. Solid phase peptide synthesis (SPPS) was used to synthesise QUB-2607, and the synthesised peptide was identified and purified by Matrix-assisted laser desorption/ionisation time of flight (MALDI-TOF) mass spectrometry (MS) and reverse phase-high performance liquid chromatography (RP-HPLC). A series of assays were performed to evaluate the bioactivity and mechanism of QUB-2607 and the results showed that QUB-2607 exhibited broad-spectrum antimicrobial activity, being especially potent against Gram-positive bacteria. However, the haemolytic activity of QUB-2607 was also strong. Furthermore, QUB-2607 could penetrate the cell membrane of bacteria according to the action mechanism assay results.

Therefore, the aim of Chapter 4 was modification of QUB-2607 to optimise its antimicrobial and haemolytic activity. A series of peptide analogues were designed by substitution of amino acids and hybridisation with another peptide, GG3. The bioactivity assay results showed that QUB-3456 exhibited broad-spectrum antimicrobial activity and weak haemolytic activity, whereas it also had a synergistic effect with conventional antibiotics and anti-infection activity in vivo. It seems that QUB-3456 combined the advantages of QUB-2607 and GG3.
In the course of searching the APD database, brevinin-1Pk was discovered, and its sequence was quite similar to that of QUB-2607 and its antibacterial activity was also broad-spectrum and especially strong against Gram-positive bacteria. Apart from this, the haemolytic activity of brevinin-1Pk was weaker than that of QUB-2607. Therefore, a series of modified peptides were designed by hybridisation of brevinin-1Pk with other chosen Rana boxes. Subsequently, the antimicrobial activities of these modified peptides were all enhanced, particularly against MRSA and the anti-biofilm actions of these peptides were dramatically increased. However, their haemolytic activities were also stronger than brevinin-1Pk, except for b-1Pk2. Among all the peptides, b-1Pk4, b-1Pk6 and QUB-G exhibited excellent antimicrobial activity, and then these peptides were evaluated for their drug potential further. These three peptides had synergistic effects with conventional antibiotics and were stable in a physiological salt environment. However, the in vivo anti-infection assay identified the cytotoxicity of these peptides which would negatively influence their clinical research. The next research step would be the optimisation of the haemolytic activity of these peptides.

Thesis is embargoed until 31 December 2027.
Date of AwardDec 2022
Original languageEnglish
Awarding Institution
  • Queen's University Belfast
SupervisorTianbao Chen (Supervisor), Mei Zhou (Supervisor) & Xiaoling Chen (Supervisor)


  • Antimicrobial peptides
  • Skin and soft tissue infections
  • Rana box
  • antimicrobial activity
  • hybridisation

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