Production and functional characterization of human insulin-like growth factor 1

Document Type: Original Research Papers

Authors

1 Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.

2 Department of Molecular Systems Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.

Abstract

Insulin-like growth factor 1 (IGF-1) is a polypeptide hormone produced mainly by the liver in response to the endocrine growth hormone (GH) stimulus. This protein is involved in a wide range of cellular functions, including cellular differentiation, transformation, apoptosis suppression, migration and cell-cycle progression and other metabolic processes. In the current study, human heart cDNA was employed to isolate IGF-1 encoding fragment using reverse transcriptase (RT) PCR. The isolated fragment was cloned into pET32a expression vector and then transformed into the competent Escherichia coli Origami 2. After selecting the correct colony with the highest expression level, the colony was cultured and induced with IPTG. Recombinant IGF-1 expression was detected by SDS-PAGE and His-tagged protein purification was performed with the affinity chromatography. In order to confirm the activity of the resultant protein, biological activity of the recombinant IGF-1 was assayed through inducing proliferation of MCF-7 cells. Molecular techniques, including PCR, restriction digestion, mass spectrometry analyses, SDS-PAGE and biological activity analyses of this protein confirmed the correct cloning, expression, and function of IGF-1 in this study. Overall, we provided a rapid and cost effective production and purification method for IGF-1 protein, which is biologically active and functional.

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