Abstract：With the promulgation of the ban on antibiotics in feed, there is an urgent need for a feed additive capable of inhibiting Gram-negative bacteria in the process of livestock and poultry breeding, so as to reduce the occurrence of diseases caused by such bacteria. The aim of this study was to design an antimicrobial peptide PC that can specifically inhibit the growth of Gram-negative bacteria, but has no significant inhibitory effect on Gram-positive bacteria. Using (RP)3WW (RP)3 as the basic unit (where P is proline, R is arginine, and W is tryptophan), the trypsin inhibitor (SFTI-1) binding loop (CTKSIPPIC) derived from sunflower (Helianthus annuus) was added to the amino terminus of the antimicrobial peptides. The secondary structure of the antimicrobial peptide PC was detected by circular dichroism (CD). And further to detect the antimicrobial activity, hemolysis, cytotoxicity and stability of the antimicrobial peptide PC. The antimicrobial peptide PC exhibited an α-helical structure under the simulated membrane hydrophobic environment. The results showed that the antimicrobial peptide PC had high inhibitory activity against Gram-negative bacteria such as Escherichia coli, Salmonella pullorum and Pseudomonas aeruginosa. In contrast, for Gram-positive bacteria such as Staphylococcus aureus, Staphylococcus epidermidis and Enterococcus faecalis, antimicrobial peptide PC did not show inhibitory activity, but could promote bacteria to produce reactive oxygen species (ROS), and had low hemolytic activity and cytotoxicity. The antimicrobial peptide PC obtained in this study had the characteristics of anti-Gram-negative bacteria, good biological safety and stability, and had the development potential to be a substitute for forage antibiotics. This study provides a basis for the design and development of antimicrobial peptides against gram bacteria..
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