噬菌体gp17基因2种重组产物的抗菌效应比较

通过重组技术获得大肠埃希菌噬菌体内溶素纯化蛋白和表面展示噬菌体,并观察产物的生物效应。将肠侵袭性大肠埃希菌EIEC 8401噬菌体LSB-1内溶素基因gp17构建到质粒pET300中,并在大肠埃希菌BL21中诱导表达,通过Ni柱纯化系统纯化产物;利用噬菌体展示技术构建T7-LSB-gp17重组噬菌体,通过双层琼脂法纯化噬菌体,并观察2种产物的抗菌效应。2 139 bp的gp17基因通过重组技术表达出78.3 ku的可溶性蛋白,纯化后浓度为2.38 mg/mL,其对EIEC8401有良好的抑菌活性,但对其他试验菌无抗性;通过噬菌体展示技术构建的重组噬菌体T7-LSB-gp17通过SDS-PAGE电泳显示在78 ku处有表达增强,对EIEC8401无感染、裂解作用,但对EIEC8401及其他试验菌有明显溶菌作用,宿主谱增加。通过重组技术获得的噬菌体LSB-1内溶素基因gp17的产物对LSB-1噬菌体原宿主具有明显的抑制效应。其中gp17表达的纯化蛋白具有明显的宿主专一性,重组噬菌体悬液有较宽种类的抗菌作用。这可能是因为gp17蛋白与噬菌体表面复杂空间结构的相互作用产生的生物效应。

Antibacterial Effect Comparison of Two Recombinant Products of Phage gp17 Gene

E.coli phage endolysin purified protein and surface display bacteriophage were obtained by recombinant technology and observe their biological effects.EIEC8401 Phage LSB-1 endolysin gene gp17 was constructed into the plasmid pET300 and induced expressed in E.coli BL21,and then the product was purified by Ni column purify system;recombinant phage T7-LSB-gp17 was constructed adopted phage display technology and purified using double layer agar method.And observe their antibacterial effects of the two products.The gp17 gene of 2 139 bp expressed a78.3 ku soluble protein through recombinant technology,the concentration was 2.38 mg /mL after purified,and had fine antibacterial activity against EIEC8401,however no resistance against other tested bacteria;the recombinant phage T7-LSB-gp17 constructed by phage display technology showed strengthened expression in 78 ku through SDSPAGE,no infection and splitting effects against,EIEC8401,but had significant bacteriolysis against EIEC8401 and other tested bacteria,the host spectrum increased.The products of phage LSB-1 endolysin gene gp17 obtained by recombinant technology had significant inhibitory effects to the original host of LSB-1 phage.Among them the purified protein had significant host specificity,and the recombinant phage suspension had a broad categories antibacterial effects.This was possibly because the surface of gp17 protein and the phage had bio-effect produced by mutual effects of complicated spatial structure.