Expression of recombinant human insulin-like growth factor type 1 [rhIGF-1] in escherichia coli

ABSTRACT

Human insulin-like growth factor type 1 [hIGF-1] is a protein consisting of 70 amino acids [MW=7.6 kDa] and mainly synthesized by liver. Mecasermin [Trade name INCRELEX] is the synthetic form of the protein which is used as an effective treatment for particular disorders such as short stature, type 1 and 2 diabetes, and wound healing. Current study was aimed to investigate the expression of human insulin-like growth factor type1 in Escherichia coli [E. coli] BL21 [DE3] expression system in order to produce an active recombinant form of the protein. For the purpose of the study, firstly codon optimization was done for hIGF-1 gene, using bioinformatics databases. Then, the gene was synthesized and inserted in pET-24a vector by a cutting strategy included NdeI and BamHI-HF enzymes. In the next step, gene was run in agarose gel and purified. The constructed expression cassette was transformed into E. coli BL21 [DE3] cells through CaCl2 heat shock method. Identification and confirmation of the transformed colonies were performed using screening PCR method. Synthesis of hIGF-1 was induced by IPTG. The expression in induced strains was analyzed by SDS-PAGE and western blotting techniques. Confirmation of cloning and IGF-1 expression cassette was carried out through genetic engineering procedures. Analysis of transformed E. coli strain with SDS-PAGE and western blotting techniques confirmed that gene was expressed in host cells. Molecular weight of the expressed protein was estimated to be 7.6 kDa. hIGF-1 expression cassette for cloning and expression in E. coli was de-signed and the protein of interest was successfully induced and identified. In addition, E. coli BL21 [DE3] can be used as a suitable host for production of recombinant hIGF-1 and this technology has a potential to be localized