Abstract
To explore the potential of heterologous protein expression in Kluyveromyces cicerisporus, three expression plasmids, pUK1-PIT, pUKD-PIT and pUKD-S-PIT, based on the vector pUK1 or pUKD were constructed and transformed, respectively, into yeast strain K. cicerisporus Y179U. Human interferon α-2a, used as an example protein, was successfully expressed and secreted by transformant Y179U/pUKD-PIT and Y179U/pUKD-S-PIT. In the flask culture, strain Y179U/pUKD-S-PIT could express interferon at 60 mg/l. The stability of plasmid pUKD-S-PIT in the host was higher than that of pUKD-PIT. This was consistent with their expression levels of interferon. There were two interferon-related bands found by Western blotting analysis. The possible reason for this is discussed.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Bianchi MM, Santarelli R, Frontali L (1991) Plasmid functions involved in the stable propagation of the pKD1 circular plasmid in Kluyveromyces lactis. Curr Genet 19:155–161
Fleer R, Chen XJ, Amellal N, Yeh P, Fournier A, Guinet F, Gault N, Faucher D, Folliard F, Fukuhara H (1991) High-level secretion of correctly processed recombinant human interleukin-1 beta in Kluyveromyces lactis. Gene 107:285–295
Hashimoto Y, Koyabu N, Imoto T (1998) Effects of signal sequences on the secretion of hen lysozyme by yeast: construction of four secretion cassette vectors. Protein Eng 11:75–77
Hitzeman RA, Leung DW, Perry LJ, Kohr WJ, Levine HL, Goeddel DV (1983) Secretion of human interferons by yeast. Science 219:620–625
Huo KK, Yu LL, Chen XJ, Li YY (1993) A stable vector for high-level expression and secretion of human interferon alpha A in yeast. Sci China B 36:557–567
Jahic M, Rotticci-Mulder JC, Martinelle M, Hult K, Enfors SO (2002) Modeling of growth and energy metabolism of Pichia pastoris producing a fusion protein. Bioprocess Biosystem Eng 24:385–393
Jayaram M, Li YY, Broach JR (1983) The yeast plasmid 2mu circle encodes components required for its high copy propagation. Cell 34:95–104
Platis D, Foster GR (2003) High yield expression, refolding, and characterization of recombinant interferon alpha2/alpha8 hybrids in Escherichia coli. Protein Expr Purif 31:222–230
Rossmann C, Sharp N, Allen G, Gewert D (1996) Expression and purification of recombinant, glycosylated human interferon alpha 2b in murine myeloma NSO cells. Protein Expr Purif 7:335–342
Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory Press, New York
Swinkels BW, van Ooyen AJ, Bonekamp FJ (1993) The yeast Kluyveromyces lactis as an efficient host for heterologous gene expression. Antonie Van Leeuwenhoek J Serol Microbiol 64:187–201
Wang H, Gao B, Chen P, Dong L, Li Y (1995) Analysis of heterogeneity of gene products (interferon) expressed in yeast. Sci China B 38:954–962
Wen TQ, Liu F, Huo KK, Li YY (2003) Cloning and analysis of the inulinase gene from Kluyeromyces cicerisporus CBS4857. World J Microbiol Biotechnol 19:426
Xu GZ, Pan F, Huo KK, Li YY (1998) Cloning of DNA Fragments Possessing Promoter Function from Kluyveromyces cicerisporus. Sheng Wu Hua Xue Yu Sheng Wu Wu Li Xue Bao (Shanghai) 30:35–40
Zhang J, Yuan H, Wen T, Xu F, Di Y, Huo K, Li YY (2003) Cloning of the KcURA3 gene and development of a transformation system for Kluyveromyces cicerisporus. Appl Microbiol Biotechnol 62:387–391
Acknowledgement
This project was granted financial support from the China Postdoctoral Science Foundation.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Cai, X.P., Zhang, J., Yuan, HY. et al. Secretory expression of heterologous protein in Kluyveromyces cicerisporus. Appl Microbiol Biotechnol 67, 364–369 (2005). https://doi.org/10.1007/s00253-004-1834-9
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00253-004-1834-9