超活性胰高血糖素的分泌表达  

Secretory Expression of the Superactive [Lys ^(17,18), Glu ^(21)]-Glucagon in E. coli

作　　者：闻崇炜[1] 甘人宝[1] 朱尚权[1] 

机构地区：[1]中国科学院上海生命科学研究院生物化学与细胞生物学研究所,上海200031

出　　处：《生物化学与生物物理学报》2003年第2期167-171,共5页

摘　　要：通过多肽化学合成方法 ,人们对胰高血糖素的结构与功能关系有了比较深刻的了解 ,其中最引人注目的成就之一是发现 [Lys17,18,Glu2 1] 胰高血糖素具有比天然胰高血糖素更高的生物活性 ,称之为超活性胰高血糖素(superactiveglucagon ,下称SA glucagon)。为了通过基因工程途径获得SA glucagon ,用PCR方法从以前构建的胰高血糖素表达载体pAGluT得到SA glucagon的基因 (SAG) ,构建了含PL 启动子 ,phoA信号肽和SAG的分泌表达载体pBLSG7。pBLSG7转化到大肠杆菌BL2 1中 ,进行SAG的分泌表达 ,在摇瓶条件下 ,该菌种能分泌表达SA glucagon达 3.6 5mg/L(A60 0 =1) ,占上清液中蛋白质的 19.5 % ,并进一步研究了诱导温度和菌株对表达的影响。One of the most important findings in structure-function studies on glucagon by means of chemical synthesis is the discovery that [Lys 17, 18, Glu 21] glucagon had higher biological activity than native glucagon. This mutant of glucagon was called superactive glucagon (SA-glucagon). In the present work, the possibility to obtain SA-glucagon by means of genetic engineering was studied. The gene of SA-glucagon (SAG) was obtained by PCR from a constructed recombinant glucagon plasmid, pAGluT. A secretory expression vector harboring SAG, pBLSG7, containing P L promoter and the gene of phoA signal peptide was constructed. In expression studies after transformation of pBLSG7 into E.coli BL21, it was found that the expression yield of SA-glucagon reached 3.65 mg/L(A 600=1), about 19.5% of total proteins in the culture medium under shaken flask conditions. In addition, the influence of induction temperature and of E.coli strain on the expression yield of SA-glucagon was also studied.

关 键 词：超活性胰高血糖素 分泌表达 基因重组 PL启动子 

分 类 号：Q786[生物学—分子生物学]