Secretory Expression of the Superactive [Lys^17,18,Glu²¹]-Glucagon in E. coli

WEN Chong-Wei, GAN Ren-Bao^*, ZHU Shang-Quan^*

( Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences,
the Chinese Academy of Sciences, Shanghai 200031, China )

Abstract One of the most important findings in structure-function studies on glucagon by means of chemical synthesis is the discovery that [Lys^17,18,Glu²¹]-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 PL 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 (A₆₀₀=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.

Key words genetical engineering; glucagon; SA-glucagon; PL promoter; secretion

Corresponding author:
ZHU Shang-Quan: Tel, 86-21-64374430-5283; Fax, 86-21-64338357; e-mail, [email protected]
GAN Ren-Bao: Tel, 86-21-64374430-5325; Fax, 86-21-64338357; e-mail, [email protected]