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Deciphering EGFP production via surface display and self-cleavage intein system in different hosts

Published on Oct 1, 2015in Journal of The Taiwan Institute of Chemical Engineers3.83
· DOI :10.1016/j.jtice.2015.03.036
Shu-Chen Kan5
Estimated H-index: 5
(NCHU: National Chung Hsing University),
Chi-Ming Chen2
Estimated H-index: 2
(NCHU: National Chung Hsing University)
+ 3 AuthorsYung-Chuan Liu24
Estimated H-index: 24
(NCHU: National Chung Hsing University)
Abstract
Abstract In this study, six E. coli strains, which harboring enhanced green fluorescence protein (EGFP), an intein (INT) and ice nucleation protein (INP), were used to express the EGFP protein. The effect of E. coli strains on the surfaced displayed EGFP via INP-INT system was first investigated. Among these strains, E. coli JM109(DE3) was found the best strain for EGFP production (67.9 mg/L), two-fold as compared to that produced by E. coli DH1(DE3). It was concluded that lon mutation could stabilize recombinant proteins. Thus, the deletion of the gene coding for proteolysis might be a critical factor when using host for surface-expression production. The optimal conditions for cell cultivation, gene induction and intein cleavage for EGFP production were systematically studied and discussed. The objective function (O) value of 26.5 can be reached at a 12-h induction and 20-h cleavage process, where the EGFP yield is 58.6 mg/L with a purity of 45.3%. Furthermore, results indicated that the higher intein activity was obtained in the shorter cleavage duration; however, the shorter cleavage time sacrificed the production yield as expected. To compromise between the production yield and purity, the optimal conditions for the EGFP production were: 12-h induction and 20-h cleavage process is for the production of higher purity EGFP.
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  • Citations (4)
References34
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#1Jiun-Yan Wu (NCHU: National Chung Hsing University)H-Index: 4
#2Tsung-Yu Tsai (NCHU: National Chung Hsing University)H-Index: 1
Last.Yung-Chuan Liu (NCHU: National Chung Hsing University)H-Index: 24
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#1Gertjan S. Verhoeven (UvA: University of Amsterdam)H-Index: 6
#2Svetlana Alexeeva (UvA: University of Amsterdam)H-Index: 11
Last.Tanneke den Blaauwen (UvA: University of Amsterdam)H-Index: 31
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