噬菌体T7溶菌酶及其融合蛋白在大肠杆菌中的表达

EXPRESSION OF T7 LYSOZYME AND ITS FUSION PROTEINSIS Escherichia coli

以噬菌体Ｔ７ＤＮＡ为模板，ＰＣＲ扩增Ｔ７溶菌酶基因，插入ｐＢｌｕｅｓｃｒｉｐｔＳＫ载体中，ＤＮＡ序列分析表明，克隆的Ｔ７溶菌酶基因和已报道的序列无氨基酸水平上的差异。将Ｔ７溶菌酶基因分别拼接在烟草病原相关蛋白（ＰＲ１ｂ）信号肽编码序列的３’末端和马铃薯卷叶病毒外壳蛋白（ＰＬＲＶＣＰ）基因靠近３’末端处，构建成两个融合蛋白基因。将Ｔ７溶菌酶及其融合蛋白基因插入大肠杆菌表达载体ｐＢＶ２２１，蛋白电泳及溶菌实验表明，Ｔ７溶菌酶基因在大肠杆菌中高效表达，其产物的表达量占菌体可溶性蛋白的２０％以上，ＰＬＲＶＣＰ的表达量并没有因Ｃ端融合Ｔ７溶菌酶而提高，高等植物的信号肽在大肠杆菌中也能起分泌信号作用。

7 lysozyme gene,which was amplified by PCR technique from bacteriophage T7 DNAtemplate, was cloned in pBluescript SK. Analysis of its nucleotide sequence showed that itsdeduced amino acid sequence was 100% homologous with those reported. T7 lysozyme genewas fused respectively to the 3′terminals of signal peptide coding sequence of tobacco PR1band potato leafroll virus coat protein gene .T7 lysozyme and its two fusion protein genes wereput in the E. coli expression vector pBV221. The yield of T7 lysozyme was more than 20%of the total bacterial soluble protein. The expression of PLRV CP was not improved through the fusion of T7 lysozyme.Signal peptide from higher plant can function as secretion signal in E.coli.