Development Of Bacterial Surface Modification Tools Based On DNA-Protein Conjugates

The dynamic changes on bacterial surface interface play a key role in interbacterial communication,bacterial survival and bacterial community.The development of suitable surface modification tools for bacteria is of great significance for understanding the communication mechanism of bacteria and studying the interaction between bacteria,host and environment.As a kind of material with programmable assembly function and high spatial precision,DNA has been widely used by researchers in the biochemical regulation of mammalian ce ll membrane interface,but the work of applying it to bacterial surface interface is still in its infancy.Nano materials on the surface of the bacteria,in order to achieve the DNA functionalized modifications,this paper developed two kinds of nucleic acid-protein conjugates tools,using protein as anchor basement,successful anchor function DNA molecules on the surface of bacteria,while the complementary base pairing function of DNA itself will not be affect,for subsequent using DNA control bacteria group behavior laid the foundation:The main research contents of this paper are as follows:(1)We have developed the first bacterial surface DNA modification tool based on a covalent conjugation of nucleic acids and lectins.We use the lectin protein(Con A),which has specific binding ability to mannose and glucose units,to recognize the high abundance of carbohydrates on the bacterial surface and to anchor DNA to the bacterial surface through the interaction.We synthesized the Con A-DNA conjugate by covalent coupling method,and characterized the purity and molecular weight of the conjugate.Subsequently,we successfully anchored DNA probes to the surface of E.coli using conjugates and demonstrated the base complementary pairing function of surface-modified DNA.The conjugate can also modify the surface of S.aureus,which proves that the method has a certain universality.(2)We developed an endonuclease Gene A* protein,which specifically recognizes and splices the properties of specific nucleic acid sequences,as the second bacterial surface DNA modification tool.Combining genetic engineering methods,we have directly demonstrated the Gene A* protein on the surface of E.coli using the Gene A*-Blc recombinant protein system.By simple co-incubation,A DNA-Gene A*conjugate can be formed on the bacterial surface to anchor DNA to the bacterial surface.We screened and optimized the high activity strains,induction culture conditions and nucleic acid incubation conditions of the genetically engineered bacteria.We proved that after induction culture,the Gene A* enzyme protein displayed on the surface of the bacteriaq with certain enzyme activity,which could specifically cut and bind DNA chains,and at the same time the surface modified DNA retained the complementary function.We successfully expressed Gene A* on the surface of three fluorescent engineered bacteria,m Cherry,GFP and BF P,and realized the formation of multi-bacterial aggregates by using the complementary base pairing function of DNA.In summary,two nucleic-protein conjugates were developed by using covalent conjugates and enzyme conjugates to achieve functional modification of DNA on bacterial surface,which is of great significance for future research on bacterial living biomaterials,bacterial population effects,and bacterial biosensors.