PRODUCTION OF BIOACTIVE GASTRIN FROM THE NONENDOCRINE CELL-LINES CHO AND COS-7

We made a mutated progastrin cDNA construct that contains a cleavage site (-Arg(-4)-Arg(-3)-Lys(-2)-Arg(-1)) specific for the Kex2-like endoprotease furin, located ahead of the bioactive gastrin. For expressing the mutated progastrin cDNA, we used two non-endocrine cell lines, CHO and COS-7. CHO cells exhibit amidating enzyme activity and levels of amidation enzyme mRNA as high as those in the pituitary-derived endocrine cell line GH,, whereas COS-7 cells have far less amidating activity and lower amounts of mRNA. Mutant progastrin-expressing CHO cells produced mostly amidated gastrin. Gel filtration showed the size of this gastrin corresponded to that of the synthetic human gastrin-17. In contrast, COS-7 cells produced glycine-extended gastrin and only a small amount of amidated gastrin. The difference in the amount of amidated gastrin products produced by the two non-endocrine cell lines is due to differing amounts of the amidation enzyme contained in each cell line.