Secretory production of a functional membrane protein

将编码来自盐生盐杆菌（Halobacterium halobium）的细菌视紫红质基因的DNA片段克隆到含有B下游的家蚕肌动蛋白启动子的质粒载体中。家蚕核型多角体病毒（BmNPV）IE-1反式激活因子和BmNPV HR 3增强子用于高水平表达。在用所得质粒转染粉纹夜蛾BTI-TN-5 B1 -4（High Five）细胞后，通过蛋白质印迹分析来分析细胞提取物。在细胞裂解物中检测到电泳迁移率约为20 kDa的特异性蛋白条带，表明细菌视紫红质在昆虫细胞中成功表达。然后将质粒载体转染到表达日本脑炎（JE）病毒膜蛋白（M）的前体（prM）和包膜糖蛋白（E）的重组High Five细胞中。已经检查了包埋细菌视紫红质的病毒样颗粒的分泌表达。

The DNA fragment encoding the bacteriorhodopsin gene from Halobacterium halobium was cloned into a plasmid vector that contained the Bombyx mori actin promoter downstream of the B. mori nucleopolyhedrovirus (BmNPV) IE-1 transactivator and the BmNPV HR3 enhancer for high-level expression. After transfection with the resultant plasmid into Trichoplusia ni BTI-TN-5B1-4 (High Five) cells, cellular extract was analyzed by western blot analysis. Specific protein bands were detected at electrophoretic mobility of approximately 20 kDa in the cell lysate, indicating successful expression of bacteriorhodopsin in insect cells. The plasmid vector was then transfected into recombinant High Five cells that expressed the precursor (prM) of the viral membrane protein (M) and the envelope glycoprotein (E) of Japanese encephalitis (JE) virus. Secretory expression of virus-like particles embedding bacteriorhodopsin has been examined.