Sato T, Matsui H, Shibahara S, Kobayashi T, Morinaga Y, Kashima N, Yamasaki S, Hamuro J, Taniguchi T
J Biochem. 1987 Feb;101(2):525-34. doi: 10.1093/oxfordjournals.jbchem.a121940.
We constructed several expression plasmids of human IL-2 gene, some of which directed high-level synthesis of mature IL-2 protein in E. coli. In all the plasmids reported here, we installed the E. coli trp promoter and SD sequence upstream of the IL-2 cDNA. When DNA sequences containing the rho-independent transcription terminator such as those involved in the trpA and lpp gene were inserted downstream of the IL-2 cDNA sequence, the expression level of the IL-2 gene increased up to 5-fold. Moreover, the deletion of either the whole region including A-T and G-C tails or a part of the 3' non-coding sequence resulted in further increase of the expression of the IL-2 gene up to 500-fold. The mature IL-2 produced in E. coli exhibited biological and immunological activities indistinguishable from those of purified IL-2 from a human T cell line, Jurkat-111. The manipulations described here may be useful for the high-level expression of eukaryotic genes in E. coli.
我们构建了几种人白细胞介素-2（IL-2）基因的表达质粒，其中一些能指导在大肠杆菌中高水平合成成熟的IL-2蛋白。在本文报道的所有质粒中，我们在IL-2 cDNA的上游安装了大肠杆菌色氨酸启动子和SD序列。当将含有不依赖于rho的转录终止子的DNA序列（如那些参与trpA和lpp基因的序列）插入IL-2 cDNA序列的下游时，IL-2基因的表达水平提高了5倍。此外，删除包括A-T和G-C尾的整个区域或部分3'非编码序列导致IL-2基因的表达进一步增加，最高可达500倍。在大肠杆菌中产生的成熟IL-2表现出与从人T细胞系Jurkat-111纯化的IL-2无法区分的生物学和免疫学活性。本文所述的操作可能有助于在大肠杆菌中高水平表达真核基因。
