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http://dx.doi.org/10.5762/KAIS.2021.22.6.542

Preparation of Recombinant Human Epidermal Growth Factor by Hydroxylamine Cleavage  

Kim, Sun-Ho (Department of Biomedical Materials, Konyang University)
Lee, Woo-Yiel (Department of Biomedical Materials, Konyang University)
Publication Information
Journal of the Korea Academia-Industrial cooperation Society / v.22, no.6, 2021 , pp. 542-549 More about this Journal
Abstract
The purpose of this study was to provide an economical and easy preparation method for recombinant human epidermal growth factor (rhEGF) without the need for an expensive enzyme to cleave the fusion part. However, the N-terminal fusion part is still useful for affinity chromatography. The hEGF is an important hormone in cell growth and proliferation in humans, and many studies on the expression and purification of this protein have been reported. In the present study, the hEGF gene was designed to be optimized with the E. coli codon usage preference and to contain Asn-Gly at the N-terminus of the protein. The gene was inserted into pRSET_A, an E. coli expression vector, and transformed into E. coli BL21 (DE3). The recombinant fusion protein was successfully co-expressed with pG-Tf2, a chaperone vector, in E. coli and purified by Ni-NTA column chromatography. The rhEGF was then released by hydroxylamine treatment and confirmed by SDS-PAGE. ELISA analysis showed that the activity of the free rhEGF was more than 92% similar to that of commercial EGF. The biological activity of the rhEGF was confirmed by a cell proliferation test with human skin fibroblasts.
Keywords
Epidermal Growth Factor; Recombination; Co-Expression; Chaperone; Hydroxylamine;
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