Production and purification of recombinant human insulin-like growth factor I from Escherichia coli

Abstract

A cDNA encoding human insulin-like growth factor I (IGF-I) was cloned and expressed in Escherichia coli. IGF-I was produced as a fusion protein with the 288 N-terminal residues of ~-galactosidase by a linker encoding hydroxylamine cleavage recognition sequence. The fusion protein was expressed with isopropyl-l-thio-beta-D-galactoside (IPTG) induction under the control of the inducible tac promotor in the form of insoluble inclusion bodies. After cleavage of the fusion protein with hydroxylamine, the released IGF-I was purified by a cation exchange chromatography, covalent chromatography, and reverse-phase high performance liquid chromatography (rp-HPLC). The identity and purity of the purified IGF-I was confirmed by N-terminal sequence analysis, SDS-polyacrylamide gel electrophoresis, and rp-HPLC. The biological activity of the purified IGF-I has been demonstrated to be indistinguishable from the native IGF-I in thymidine uptake, protein synthesis and radioreceptor assay.