Recombinant Expression And Bioactivity Of Antimicrobial Peptides

In recent years, the abuse of antibiotics accelerated the antibiotic resistance evolution of pathogenic strains. The resistance of bacterial has became a global tissue.So it’s urgent to find new drugs with structural uniqueness to instead of the traditional antibiotics.The antimicrobial mechanism of antimicrobial peptides(AMPs) is different with antibiotics, AMPs are environmentally friendly, so it became a new type of antibacterial drug and additive..In this study, the anti-lipopolysaccharide factor 3 from Penaeus monodon was investigated as antimicrobial peptide to against microorganisms. The plasmid pET22b-ALFpm3 with soluble expression and plasmid pET22b-ALFpm3-PT18 A with self-assembled short peptide fusion expression were constructed. In antimicrobial activity assay, the recombinant ALFpm3 inhibited the growth of E.coli DH5α, B.subtilis 168 and S.aureus, and the inhibition zone of E.coli DH5α,. B.subtilis168 and S.aureus was about 25 mm, 23 mm and 13 mm, respectively. However, the recombinant ALFpm3-PT18 A didn’t show antimicrobial activity. The concentration of purified recombinant ALFpm3 was 23 μg/mL. In thermostability test, ALFpm3 had an amazing performance, the antibacterial activity of which didn’t reduce after boiling 3h. But the yield of recombinant ALFpm3 in E.coli was too low to large scale of preparation..In order to promote the production of AMPs, the secretion expression plasmid pPIC9K-ALFpm3 was constructed and transformed into P.pastoris KM71. The ALFpm3 could be detected By Western Blot in shaking flask fermentation. After high density fermentation, the yield of ALFpm3 was 75 μg/ml, but almost have no antibacterial activity.This study demonstrated that ALFpm3 has a potential application. Unfortunately, this study failed to got large amount of ALFpm3, if you want to study its activity in further, you can try other host and plasmid.