Novel secretion system of recombinant Saccharomyces cerevisiae using an N-terminus residue of human IL-1β as secretion enhancer

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Title
Novel secretion system of recombinant Saccharomyces cerevisiae using an N-terminus residue of human IL-1β as secretion enhancer
Author(s)
Jee Won Lee; Seong Il Choi; Jun Sung Jang; Ki Ryong Jang; Jae Woong Moon; Cheon Soon Bae; Doo Suk Yang; Baik Lin Seong
Bibliographic Citation
Biotechnology Progress, vol. 15, pp. 884-890
Publication Year
1999
Abstract
An N-terminus sequence of human interleukin 1β (hIL-1β) was used as a fusion expression partner for the production of two recombinant therapeutic proteins, human granulocyte-colony stimulating factor (hG-CSF) and human growth hormone (hGH), using Saccharomyces cerevisiae as a host. The expression cassette comprised the leader sequence of killer toxin of Kluyveromyces lactis, the N-terminus 24 amino acids (Ser5-Ala28) of mature hIL-1β, the KEX2 dibasic endopeptidase cleavage site, and the target protein (hG-CSF or hGH). The gene expression was controlled by the inducible UAS(gal)/MF-α1 promoter. With the expression vector above, both recombinant proteins were well secreted into culture medium with high secretion efficiencies, and especially, the recombinant hGH was accumulated up to around 1.3 g/L in the culture broth. This is due presumably to the significant role of fused hIL-1β as secretion enhancer in the yeast secretory pathway. In our recent report, various immunoblotting analyses have shown that the presence of a core N-glycosylation resident in the hIL-1β fragment is likely to be of crucial importance in the high-level secretion of hG-CSF from the recombinant S. cerevisiae. When the N-glycosylation was completely blocked with the addition of tunicamycin to the culture, the secretion of hG-CSF and hGH was decreased to a negligible level although the other host-derived proteins were well secreted to the culture broth regardless of the presence of tunicamycin. The N-terminal sequencing of the purified hG-CSF verified that the hIL-1β fusion peptide was correctly removed by in vivo KEX2 protease upon the exit of fusion protein from Golgi complex. From the results presented in this article, it is strongly suggested that the N-terminus fusion of the hIL-1β peptide could be utilized as a potent secretion enhancer in the expression systems designed for the secretory production of other heterologous proteins from S. cerevisiae.
ISSN
8756-7938
Publisher
Wiley
DOI
http://dx.doi.org/10.1021/bp9900918
Type
Article
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1. Journal Articles > Journal Articles
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