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Deletion Analysis of Pichia PGK1 Promoter and Construction of an Episomal Vector for Heterologous Protein Expression in P. pastoris  

Lee, Sung-Jae (Department of Bioengineering and Technology, Kangwon National University)
Hong, In-Pyo (Department of Bioengineering and Technology, Kangwon National University)
Baek, Seon-Yeol (Department of Bioengineering and Technology, Kangwon National University)
Choi, Shin-Geon (Department of Bioengineering and Technology, Kangwon National University)
Publication Information
Microbiology and Biotechnology Letters / v.35, no.3, 2007 , pp. 184-190 More about this Journal
Abstract
Approximately 2.0 kb of the promoter region of the Pichia pastoris phosphoglycerate kinase gene (PGK1) was reduced to a 266 bp fragment and this minimized portion was used for construction of a new episomal constitutive expression vector in P. pastoris. As an approach to developing a constitutive expression vector in P. pastoris, the GAP promoter region of the Pichia expression vector pGAPZB was replaced with sequentially deleted PGK1 promoter fragments fused to a beta-galactosidase gene. When a lacZ gene was used as a reporter gene, PGK1 promoter strength was lower than that of the constitutive GAP promoter but it was higher than TEF1. We report here the development of the pPGKZ-E vector as a new episomal expression vector for heterologous gene expression by removing non-essential regions of the PGK1 promoter. This broadens the choice of episomal expression vectors for controlled constitutive expression in P. pastoris.
Keywords
Pichia pastoris; PGK1; promoter; expression vector;
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1 Almeida, de J.R.M., de L.M.P. Moraes, and F.A.G Torres. 2005 Molecular characterization of the 3-phosphoglycerate kinase gene (PGK1) from the methylotrophic yeast Pichia pastoris. Yeast 22: 725-737   DOI   ScienceOn
2 Mumberg, D., R. Muller, and M. Funk. 1995. Yeast vectors for the controlled expression of heterologous proteins in different genetic backgrounds. Gene 156: 119-122   DOI   ScienceOn
3 Nacken, V., TAchstetter, E. Degryse. 1996. Probing the limits of expression levels by varying promoter strength and plasmid copy number in Saccharomyces cerevisiae. Gene 175: 253-260   DOI   ScienceOn
4 Sears, I., J. O'Connor, O. Rossanese, and B.Glick. 1998. A versatile set of vectors for constitutive and regulated gene expression in Pichia pastoris. Yeast 14: 783-790   DOI   ScienceOn
5 Gellissen, G, G Kunze, C. Gaillardin, lM. Cregg, E. Berardi, M.Veenhuis, and J. Klei. 2005. New yeast expression platforms based on methylotrophic Hansenular polymorpha and Pichia pastoris and on dimorphic Arxula adeninivorans and Yarrovwia lipolytica- A comparison. FEMS Yeast Research 5: 1079-1096   DOI   ScienceOn
6 Hong, J.P., S.J. Lee, Y.S. Kim, and S.G Choi. 2007. Recombinant expression of human cathelicidin (hCAP/LL37) in Pichia pastoris. Biotechol. Lett. 29: 73-78
7 Menendez, J, L. Hernadez, A., Banguela, and J. Pais. 2004. Functional production and secretion of the Gluconoacetobacter diazatrophicus fructose-relating exo-levanase(LsdB) in Pichia pastoris. Enz. Microb. Technol. 34: 446-452   DOI   ScienceOn
8 Hong, J.P., S. Anderson, and S.G Choi. 2006. Evaluation of a new episomal vector based on the GAP promoter for structural genomics in Pichia pastoris. J. Microbiol. Biotechnol. 16: 1362-1368   과학기술학회마을
9 Macauley-Patrick, S., M. L. Fazenda, R. McNeil, and L.M. Harvey. 2005 Heterologous protein expression using the Pichia pastoris expression system. Yeast 22: 249-270   DOI   ScienceOn
10 Lee, C.C., T.G Williams, D. Wong, and GH. Robertson. 2004. An episomal expression vector for screening mutant gene libraries in Pichia pastoris. Plasmid 54: 80-85   DOI   ScienceOn
11 Monfort, A., S.,Finger, P. Sanz, and J.A. Prieto. 1999. Evaluation of different promoters for the efficient production of heterolgous proteins in baker's yeast. Biotechnol. Lett. 21: 225-229   DOI   ScienceOn
12 Lueking, A., C. Holz, C. Gotthold, H. Lehrach, and D. Cahill. 2000. A system for dual protein expression in Pichia pastoris. Protein. Express. Purif. 20: 373-378
13 Cregg, J.M., K.J. Barringer, A.Y.Hessler, and K.R. Madden. 1985. Pichia pastoris as a host system for transformations. Mol. Cell. Biol. 5: 3376-3385   DOI   PUBMED
14 Ahn, J.H., J.Y. Hong, H.K. Lee, M.S. Park, E.G Lee, C.S. Kim, E.S. Choi, J.K. Jung, and H.W. Lee. 2007. Translation elongation factor-I gene from Pichia pastoris: molecular cloning, sequence, and use of its promoter. Appl. Microbiol. Biotechnol. 74: 601-608   DOI
15 Tschopp, J., P. Brust, J. Cregg, C. Stillman, and T. Gingeras. 1987. Expression of the lacZ gene from two methanol-regulated promoters in Pichia pastoris. Nucleic Acids Res. 15: 3859-3876   DOI   ScienceOn
16 Chung, K.S., H.S. Kang, K.W. Kim, I. Choi, K.H. Pyun, and H.S. Yoo, H.S. 1997. Expression of recombinant human interleukin (rhIL6) in Saccharomyces cerevisiae by the modified phosphoglycerate kinase and chelatin promoter. Biotechol. Lett. 19: 1169-1173   DOI   ScienceOn
17 Sambrook, J., and D.W. Russell. 2001. Molecular cloning-A laboratory manual. 3rd Ed. CSHL Press, Cold Spring Harbor, New York