Rational design and engineering of antimicrobial peptides found from natural sources

Nature, the treasure house of humankind, has created and possesses almost inexhaustible natural sources. It is an infinite treasure trove of drug discovery and research, as well as a stent for effective drugs to expand a variety of bioactive agents that are valuable for diseases or symptoms. This project aims to isolate and identify novel AMPs from Nature with potent biological effects and investigate the structure activity relationships of these peptides in addition. In Chapter 3, a 13 amino acid peptide, crabrolin, has been evaluated for biological activity, and seven derivatives were designed by changing the number, type and distribution of its charged residues and these were expected to produce changes in its biological activity. The results demonstrated that crabrolin had moderate antibacterial and anticancer activity. However, its designed analogues showed antibacterial and anti-cancer effects superior to those of the parent peptide, especially crabrolin-TR, which carries four positively charged amino acids and which had the highest activities on tested bacteria and cancer cells. These results suggested that a reasonable increase in the number and distribution of charges could raise the anti-bacterial and anti-cancer activities of crabrolin. Crabrolin-TR has the potential to be a new anti-bacterial and anti-cancer dual effect drug due to its outstanding performance in killing drug-resistant bacteria and cancer cell lines. In Chapter 4, GV 30 was successfully identified in the skin secretion of Gunther's frog (Hylarana guentheri) for the first time using "shotgun" cloning. GV 30 was proven to have potent broad-spectrum antibacterial activity, along with a low haemolytic effect. In addition, GV 30 also had a significant anti-proliferative activity against five tested cancer cell lines and produced less damage to normal human cells. In addition, GV30 showed a significant synergistic effect with mitomycin and taxol with weaker toxicity to healthy cells compared with using single drugs. Also, GV30 was also proven to cause apoptosis at a lower treatment dose through cell cycle arrest and through necrosis at a higher dose by destroying the integrity of the cell membrane. In summary, GV 30 was discovered in frog secretion and has the potential to become an anticancer drug due to its significant anti-proliferation effects and additional synergistic effects with specific chemotherapy drugs. In Chapter 5, seven truncated derivatives of GV 30 were designed based on the prediction of cleavage after treatment with trypsin and these peptides were tested for antibacterial effects, anti-biofilm ability, antibacterial kinetics, anti-proliferation, toxicity and salt stability. Among these, GV 21 (GVIFNALKGVAKTVAAQLLKK-NH2) had development potential as a new MRSA specific antibacterial agent because of its faster antibacterial effect against MRSA strains in vivo and in vitro, and its lower cytotoxicity.