Study Of Proteins Decorated With Antibacterial Polymers By In-situ Polymerization

Most of the antibacterial proteins have a certain role in inhibiting bacteria or fungi and have been successfully applied to the control of animal and plant pathogens.However,the antibacterial protein has the problems of narrow antibacterial range,easy deactivation,and vulnerability to environmental changes,which limits its development in the field of food,medicine and health.The antibacterial polymer has a good antibacterial effect,a wide range of antibacterial properties,easy availability of raw materials,and good tolerance,is a stable organic antibacterial agent.This study intends to combine the advantages of two antibacterial materials to form an antibacterial protein polymer complex to improve the performance of antibacterial materials.Antimicrobial polymers were grafted from antimicrobial proteins,and a new research method and ideas for using chemical modification synthetic methods to combine biomacromolecules like proteins with organic antimicrobial polymers were explored.The Bacillus amyloliquefaciens NK-2 natural strain secretes yhfE protein with bacteriostatic action,clones yhfE gene from the genome of Bacillus amyloliquefaciens MG3 strain,then constructs PET-28a-yhfE recombinant expression vector and expression in E.coli BL21 by Escherichia coli prokaryotic expression System.After the cell disruption,the protein was purified by a nickel ion affinity chr-omatography column to obtain yhfE target protein with a molecular weight of about 46KDa.The yhfE protein was modified with pyridoxal-5-phosphate(PLP)in a certain ratio to obtain the yhfE-CHO protein whose N-terminus was modified to an aldehyde group.The yhfE-i was obtained after reacting the yhfE-CHO protein with the synthetic initiator(2-(aminooxy)ethyl)2-bromo-2-methylpropanoate(ABM)at a certain molar ratio.The protein yhfE-i were surface-initiated polymerized with the antiadhesive monomer Poly(ethylene glycol)methacrylate(OEGMA)and the antibacterial monomer Methacroylcholine Chloride(MTAC)to obtain the protein-polymer products yhfE-OEGMA,yhfE-OEGMA&MAC,and yhfE-MTAC.The results of protein electrophoresis and nanoparticle size analyzers indicated that the surface of the protein was successfully modified with a macromolecule.Antimicrobial experiments showed that the yhfE protein has a weak bacteriostatic ability against the fungus loquat Coctobacillus,and almost no bacteriostatic effect against bacteria,including gram-negative bacteria and positive bacteria.The results of turbidity experiment,platelet inhibition zone experiment and slide protein imprinting experiment showed that the protein of the polymeric modified antibacterial polymer has obvious inhibitory effect on the bacteria,and the antibacterial effect on the fungus is also enhanced.In this study,we successfully expressed and purified yhfE antibacterial protein and synthesize antibacterial protein polymer complexes by surface-initiated polymerization to verify the antibacterial activity.This study provides ideas and preliminary verification for the development of new broad-spectrum and high-efficiency antibacterial materials.