Cloning, Expression and Activity Analysis of Avian β-Defensin 6 and 9

PurposeTo explore the prokaryotic expression and its biological characteristics of recombinant avian defensins 6 (AvBD6) and avian defensins 9 (AvBD9).

MethodThe mature peptide genes of AvBD6 and AvBD9 were cloned into the expression vector pgex-6p-1, and the recombinant expression plasmids pgex-6p-1-AvBD6 and pgex-6p-1-AvBD9 were constructed. Then the recombinant expression plasmid was transformed into BL21(DE3)pLysS. Purification after IPTG induced protein expression SDS-PAGE was used to analyze the protein expression of AvBD6 and AvBD9, then to detect their antibacterial activity in vitro and to observe the morphological changes of bacteria with transmission electron microscope. In addition, the hemolytic activity of recombinant protein on chicken erythrocytes was detected.

ResultsBy SDS-PAGE analysis, the molecular weight of the induced AvBD6 and AvBD9 is about 30 ku, and it was mainly expressed in the supernatant. The recombinant protein had antibacterial activity against Escherichia coli and Staphylococcus aureus. Electron microscopic observation showed that the bacterial contents leaked out and the damage was serious, and the hemolytic activity of the recombinant protein to chicken red blood cells was very low.

ConclusionThe recombinant protein of AvBD6 and AvBD9 prepared in this study has good biological safety and antibacterial effect, and laid a foundation for further study of its antibacterial mechanism and preliminary clinical application.