Keywords

Antibacterial Protein, Fusion Protein, Bactericidal/Permeability Increasing Protein, Ll, Antibacterial, Sepsis

Abstract

Objective To construct a fusion protein (BPI-LL37) expression vector of bactericidal/permeability increasing protein and LL37 antibacterial protein, obtain a multi-effect antibacterial fusion protein for sepsis treatment, and provide new measures for sepsis treatment. Methods The required active fragments of rBPI21 and LL37 were amplified and modified from the cDNA of bactericidal/permeability-increasing protein (BPI) and LL37 genes through PCR method, and the rBPI21-LL37/LL37-rBPI21 fusion protein was constructed. Expression vector, the BPI and LL37 genes on the constructed expression vector (pLVX-Tight-Puro) are connected through a flexible segment (GGSGG). PolyJetTM in vitro DNA transfection reagent was then used to transfect human lung adenocarcinoma cells A549. Westernblot was used to detect the level of the fusion protein expressed in eukaryotic cells A549, and the antibacterial test was used to verify the secretion and antibacterial ability of the fusion protein. Results: Restriction enzyme digestion, DNA electrophoresis and DNA sequencing proved that the fusion protein expression vector (pLVX-Tight-Puro-BPI-LL-37) was successfully constructed; Westernblot detection results showed that the fusion protein rBPI21-LL37/LL37-rBPI21 could be Expressed in eukaryotic A549 cells, the size of the fusion protein ranges from 25 to 30¡Á103; the antibacterial test further verified that the fusion protein has the biological effect of inhibiting bacterial growth. Conclusion The two constructed rBPI21-LL37/LL37-rBPI21 expression vectors can be successfully transfected into human somatic cells, and express and secrete fusion proteins with antibacterial activity. This fusion protein has the potential to become a new drug for the treatment of sepsis.

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Original research done by Yuan Peisong, Zhu Ming, Guo, Xing Wei, Li Xiangyun, He Jing, Liang Huaping, Jiang Dongpo, Xu Xiang, Huang Hong

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