Effects of L-arginine on refolding of lysine-tagged human insulin-like growth factor 1 expressed in Escherichia coli
- Authors
- Choi, Seung Phill; Park, Yong-Cheol; Lee, JungHwa; Sim, Sang Jun; Chang, Ho-Nam
- Issue Date
- 1월-2012
- Publisher
- SPRINGER
- Keywords
- Insulin-like growth factor 1; Inclusion body; Refolding; L-arginine
- Citation
- BIOPROCESS AND BIOSYSTEMS ENGINEERING, v.35, no.1-2, pp.255 - 263
- Indexed
- SCIE
SCOPUS
- Journal Title
- BIOPROCESS AND BIOSYSTEMS ENGINEERING
- Volume
- 35
- Number
- 1-2
- Start Page
- 255
- End Page
- 263
- URI
- https://scholar.korea.ac.kr/handle/2021.sw.korea/109183
- DOI
- 10.1007/s00449-011-0619-7
- ISSN
- 1615-7591
- Abstract
- Insulin-like growth factor 1 (IGF1), a therapeutic protein, is highly homologous to proinsulin in 3-dimensional structure. To highly express IGF1 in recombinant Escherichia coli, IGF1 was engineered to be fused with the 6-lysine tag and ubiquitin at its N-terminus (K6Ub-IGF1). Fed-batch fermentation of E. coli TG1/pAPT-K6Ub-IGF1 resulted in 60.8 g/L of dry cell mass, 18% of which was inclusion bodies composed of K6Ub-IGF1. Subsequent refolding processes were conducted using accumulated inclusion bodies. An environment of 50 mM bicine buffer (pH 8.5), 125 mM l-arginine, and 4 A degrees C was chosen to optimize the refolding of K6Ub-IGF1, and 240 mg/L of denatured K6Ub-IGF1 was refolded with a 32% yield. The positive effect of l-arginine on K6Ub-IGF1 refolding might be ascribed to preventing unfolded K6Ub-IGF1 from undergoing self-aggregation and thus increasing its solubility. The simple dilution refolding, followed by cleavage of the fusion protein by site-specific UBP1 and chromatographic purification of IGF1, led production of authentic IGF1 with 97% purity and an 8.5% purification yield, starting from 500 mg of inclusion bodies composed of K6Ub-IGF1, as verified by various analytical tools, such as RP-HPLC, CD spectroscopy, MALDI-TOF mass spectrometry, and Western blotting. Thus, it was confirmed that l-arginine with an aggregation-protecting ability could be applied to the development of refolding processes for other inclusion body-derived proteins.
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Collections - College of Engineering > Department of Chemical and Biological Engineering > 1. Journal Articles
- College of Medicine > Department of Medical Science > 1. Journal Articles
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