Development of HIV-1 protease expression methods using the T7 phage promoter system
Development of HIV-1 protease expression methods using the T7 phage promoter system
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DOI：
10.1007/s002530050920
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发表时间：
1997-03-01
影响因子：
5
通讯作者：
Handa, H
中科院分区：
文献类型：
--
作者：
Komai, T;Ishikawa, Y;Handa, H
利用T7噬菌体启动子系统，建立了人类免疫缺陷病毒1型(HIV-1)蛋白水解酶在大肠杆菌中的表达方法。为了在T7聚合酶的控制下抑制HIV-1蛋白的表达，我们测试了两种新方法。其中一个涉及在T7启动子的控制下将补充的T7启动子区域导入含有HIV-1蛋白酶基因的宿主细胞[E.ColiBL-21(DE3)]。预计补充的T7启动子区域将与HIV-1蛋白酶表达载体竞争T7聚合酶结合。另一起涉及携带与表达T7聚合酶的M13噬菌体相同的HIV-1蛋白酶表达载体的大肠杆菌JM109的对数晚期培养物的感染。这两种方法都是有效的，并且用成熟的HIV-1蛋白酶表达载体获得的转化子的HIV-1蛋白酶活性比用自动处理载体获得的活性高10倍。所述的表达系统不仅方便，而且很容易应用于其他对大肠杆菌有毒的蛋白的表达。
New and simple human immunodeficiency virus type 1 (HIV-1) protease expression methods in Escherichia coli were developed using the T7 phage promoter system. In order to suppress leaky HIV-1 protease expression under the control of the T7 polymerase, two new methods were tested. One involved the introduction of supplementary T7 promoter regions into host cells [E. coli BL-21(DE3)] containing the HIV-1 protease gene under the control of the T7 promoter. It was expected that the supplementary T7 promoter regions would compete with the HIV-1 protease expression vector for the T7 polymerase binding. The other involved the infection of late-log-phase cultures of E. coli JM109 harboring the same HIV-1 protease expression vector with the M13 phage expressing T7 polymerase. Both methods were effective, and transformants with the mature HIV-1 protease expression vector showed ten times higher HIV-1 protease activity than activities obtained with the autoprocessing vector. The expression systems described here are convenient and are also easily applicable for the expression of other proteins toxic for E. coli.