EXPRESSION OF RECOMBINANT HUMAN INSULIN-LIKE GROWTH FACTOR-I IN MAMMALIAN-CELLS

被引：10

作者：

BOVENBERG, WA

DAUWERSE, JG

POSPIECH, HM

VANBUULOFFERS, SC

VANDENBRANDE, JL

SUSSENBACH, JS

机构：

[1] STATE UNIV UTRECHT,PHYSIOL CHEM LAB,VONDELLAAN 24A,3521 GG UTRECHT,NETHERLANDS

[2] STATE UNIV UTRECHT,INST MOLEC BIOL & MED BIOTECHNOL,UTRECHT,NETHERLANDS

[3] STATE UNIV UTRECHT,DEPT PEDIAT ENDOCRINOL,UTRECHT,NETHERLANDS

来源：

MOLECULAR AND CELLULAR ENDOCRINOLOGY | 1990年 / 74卷 / 01期

关键词：

Expression system; Expression vector; Insulin-like growth factor I; Mammalian cells; Recombinant insulin-like growth factor I; Transfection;

D O I：

10.1016/0303-7207(90)90204-L

中图分类号：

Q2 [细胞生物学];

学科分类号：

071009 ; 090102 ;

摘要：

In order to develop a suitable mammalian expression system for human insulin-like growth factors (hIGFs) and mutant IGFs, we have constructed several artificial IGF genes, based on a cDNA encoding the IGF-I precursor (153 amino acids). Transient expression experiments using mouse Ltk- cells revealed that the IGF-I gene constructs were efficiently expressed when placed under control of the SV40 Early promoter (SV40E). This resulted in the synthesis and secretion of IGF-I receptor-reactive products. Constructs encoding an IGF-I precursor with a truncated signal peptide of 25 amino acids under control of SV40E promoter or the inducible Drosophila heat shock hsp70 promoter, were used to establish stably transformed CHOdhfr- and mouse L cells. Clones secreting IGF-I were identified by an IGF-I-specific radioreceptor assay. Immunoblot analysis of conditioned media from these clones resulted in the specific precipitation of a protein of 7 kDa identical in size to native IGF-I purified from human serum. After optimization of the expression conditions, the stable cell lines secrete 0.5-2 μg/106 cells of IGF-I. The biological activity of the secreted recombinant IGF-I was shown by its ability to stimulate DNA synthesis in human MCF-7 cells. The results described in this paper indicate that a mammalian expression system, employing CHOdhfr- or L cells, is a useful system for the synthesis of biological active IGF-I. © 1990.

引用

页码：45 / 59

页数：15

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