Virtual Screening,activity Research And Application In Chilled Meat Of Multi-target Antibacterial Lead Compound

Pseudomonas aeruginosa(P.aeruginosa)and Escherichia coli(E.coli)are the common pathogenic bacteria in food and hospital,widely exist in the daily life,and whose resistance to P-lactam antibiotics has brought great challenges to human health and medical development.The penicillin-binding protein 3 of P.aeruginosa and penicillin-binding protein 1b,4 and 5 of E.coli are play an important role in the synthesis of the cell wall,the growth of bacteria and maintaining cell morphology.In this paper,the four kinds of PBPs were selected as the targets,molecular docking and virtual screening were used to find a lead compound,which is not only can effectively inhibit the growth of bacteria,but whose structure without β-lactam ring to help to solve the problem of drug resistance of bacteria.In addition,the lead compound which acts on the cell wall of bacteria to exert antimicrobial effect was used as food preservative to verify the preservation effect on chilled meat during storage.The main contents and conclusions as follows:1.The DOCK6.5 was selected and all the compounds in the ZINC database were used as ligands,whose conformation were adjusted and projected into the active pockets of receptor proteins of penicillin-binding protein 3 of P.aeruginosa and penicillin-binding protein 1b,4,5 of E.coli to calculated the interaction energy.After the first round of virtual screening,all the compounds whose Grid score below-30 kal/mol were selected into the second round,and ligands of Amber score lower than-20 kal/mol were obtained.Finally,after the analysis of these ligands,the compound of ZINC00053282 was selected for follow-up research.2.According to the results of virtual screening,the 2,4,6-Trichloro-1,3,5-triazine(TCT)was used as raw material,N,N-Diisopropylethylamine(DIPEA)and sodium hydroxide(NaOH)were selected as acid binding agent,three chlorine of TCT were substituted by piperidinyl and two glycine molecules to synthetise the lead compound,which purified by silica gel column and the structure was verified by nuclear magnetic resonance and mass spectrometry.3.The broth dilution method was used to determine the minimum inhibitory concentration(MIC)of lead compound for five kinds of bacteria.The results showed that the MIC of ZINC00053282 to P.aeruginosa,E.coli,B.subtilis,L.monocytogenes and S.aureus were 200μg/mL,175 μg/mL,225 μg/mL,225 μg/mL and 250 μg/mL,which proved the lead compound has antibacterial activity for both Gram-positive and Gram-negative bacteria,and better than tea polyhenols(800-1600 μg/mL)which is used as preservative regularly in food,but it worse than ampicillin sodium(12.5 μg/mL).4.The color,the total number of colonies,pH,TVB-N value and meat quality loss rate of chilled meat were determined to test the preservation capability of lead compound,and set the potassium sorbate as the control group.The results showed that there were significant differences between lead compound and the blank control group on five indicators(P<0.05).On the other hand,the lead compound were better than potassium sorbate in control the increase of pH and TVB-N value,reduce the drip loss,and inhibit the growth of bacteria.Besides,compared with two groups,the lead compound has a certain function of color protection,but whose ability of it was weaker than potassium sorbate.In a word,the lead compound could effectively delay the spoilage of chilled meat,prolong the shelf life of 3～5 d.