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KMID : 1094720090140020161
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Biotechnology and Bioprocess Engineering
2009 Volume.14 No. 2 p.161 ~ p.167
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Stable Constitution of Artificial Oil Body for the Refolding of IGF1
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Choi Seung-Phill
Chang Ho-Nam
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Abstract
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Over-expression of oleosin-fused IGF1 results in the formation of insoluble aggregates in Escherichia coli occupying 35긯 of total proteins. In this study, a method based on artificial oil body (AOB) was applied to obtain active IGF1, insulin-like growth factor 1, from its insoluble form in one step. The stability of AOB emulsions constituted with soybean oleosin was maximized in the optimal composition of TAG (97.04%, wt/wt), PL (1.14%, wt/wt), and oleosin-UbIGF1 (1.82%, wt/wt) at pH 7.5 and at 25꼆C. Upon sonication, the mixture comprising plant oil and the insoluble fusion protein was readily assembled into AOBs. After peptide cleavage mediated by endopeptidase, the IGF1 free of fusion tags was liberated and then recovered. Subsequently, IGF1 self-refolded on AOB was obtained with high yield of 63.2 mg/g dry cell. This on-AOB refolding can be applied to the development of bacterial expression and purification of other active recombinant proteins.
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KEYWORD
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insulin-like growth factor, artificial oil body, oleosin, refolding
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