Soluble expression and activity evaluation of antibacterial peptide-endolysin fusion anti-pseudomonas aeruginosa protein
OBJECTIVE To express and purify an antibacterial peptide-endolysin fusion protein by fusion expression with Trigger Factor,and initially evaluated its antibacterial activity.METHODS A new antibacterial protein AL was designed by fusing 29 amino acids at the N-terminus of the sheep myeloid antibacterial peptide with the C-terminus of endolysin LysPA26.pCold-His-TF-AL expression plasmid was constructed by homologous recombination.Antibacterial protein AL was purified by Ni-affinity chromatography and anion-exchange chromatography,and the anti-Pseudomonas aeruginosa activity of AL was initially evaluated by plate counting method.RESULTS In E.coli BL21(DE3),the plasmid pCold-His-TF-AL could significantly express the antibacterial protein AL,and more than 90％of AL was distributed in the supernatant of E.coli lysate in the soluble form.The anti-Pseudomonas aeruginosa activity of AL was evaluated by plate counting method,which showed that 0.05 mg·mL-1 AL could reduce the amount of P.aeruginosa PAO1 by 5.55 log units within 30 min,with high antibacterial activity.CONCLUSION Efficient soluble expression of AL can be achieved by using Tigger Factor fusion expression and cold shock induction method,and the purified AL had significant antibacterial activity against P.aeruginosa.
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