Abstract
We have recently employed an intein, Saccharomyces cerevisiae vascular membrane ATPase (VMA), in conjunction with efficient expression and secretory functions formed between the ompA leader sequence and the human epidermal growth factor (EGF) gene (fused at the 5′ end of VMA), and the human basic fibroblast growth factor (bFGF) gene (fused at the 3′ end of VMA), to engineer an efficient intein-based Escherichia coli system for high-level co-expression of EGF and bFGF as authentic mature products. Both products were found not only excreted to the culture medium but also located, surprisingly, in the cytoplasm (Kwong and Wong 2013). In this study, we employed two structurally varied inteins, VMA and Mycobacterium xenopi GyraseA (GyrA), and further demonstrated that despite acting alone, both VMA and GyrA were able to mediate successful co-expression of two widely different proteins, EGF and an endoglucanase (Eng) in E. coli. Although EGF and Eng were initially expressed as large precursors/intermediates, they were soluble and auto-cleavable to finally yield the desired products in both the cytoplasm and culture media. The results further substantiate our postulation that the aforementioned intein/E. coli approach might lead to the development of cost-effective and versatile host systems, wherein all culture fractions are involved in producing the target proteins.
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Ethical statement
This study was funded by RGC project: GRF16101515, and Research Contracts No. BWT001.04/05 and GVN002.06/07 managed by Research and Development of HKUST. The three authors, Keith W. Y. Kwong, Alan, K. L. Ng, and W.K.R. Wong, declare that they have no conflict of interest. This article does not contain any studies with human participants or animals performed by any of the authors.
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Kwong, K.W.Y., Ng, A.K.L. & Wong, W.K.R. Engineering versatile protein expression systems mediated by inteins in Escherichia coli . Appl Microbiol Biotechnol 100, 255–262 (2016). https://doi.org/10.1007/s00253-015-6960-z
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DOI: https://doi.org/10.1007/s00253-015-6960-z