Prokaryotic Expression and Functional Verification of Antimicrobial Peptide LR_GG

The antimicrobial peptide LR_GG (LLRLLRRGGRRLLRLL-NH2) was designed and chemically synthesized in a study conducted by Jia et al. Gram-negative bacteria were found to be sensitive to LR_GG and exhibited a high therapeutic index. Genetic engineering methods were used to create the prokaryotic fusion expression vector pQE-GFP-LR_GG, and the resulting corresponding fusion protein GFP-LR_GG was subsequently expressed and purified. The precursor GFP was then removed by TEV proteolysis, and pure LR_GG was obtained after another round of purification and endotoxin removal. The prokaryotic-expressed antimicrobial peptide LR_GG displays a broad-spectrum antibacterial effect on Gram-negative bacteria, and its minimum inhibitory activity (MIC) against Escherichia coli can reach 2 μg/mL. Compared to the chemically synthesized LR_GG, the prokaryotic-expressed LR_GG exhibits similar temperature, pH, salt ion, serum stability, and cell selectivity. Furthermore, prokaryotic-expressed LR_GG showed excellent therapeutic effects in both the infection model of cell selectivity and no embryotoxicity in a Galleria mellonella infection model. The mechanism by which LR_GG causes bacterial death was found to be the disruption of the Gram-negative cell membrane.