Abstract
In this study, we designed an expression system for the Y. lipolytica yeast, which can be fully efficient in media with non-standard industrial ingredients. Previously, we reported that the mitochondrial Voltage Dependent Anion Channel (VDAC) was a major protein overproduced in the Yarrowia lipolytica yeast in an alkaline (pH = 9.0) culture medium. In this study, the VDAC promoter was cloned and tested using a reporter system based on the LacZ gene. Naturally, the VDAC gene contains an intron located just within the ATG translation initiation codon. The VDAC promoter V2 variant with the intron and V3 variant without the intron were studied. The VDAC-driven clones exhibited high variability of the expression profile. Some clones were more active than the clones induced by the artificial hp4d promoter, when grown in both complete and low-cost industrial ingredient (10% fish and sunflower meal) containing media. The new expression system may greatly expand the range of recombinant producers of feed enzymes and some other types of fodder additives in the Y. lipolytica yeast, appropriate for assimilation of low-cost and non-standard raw material.
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References
Barth, G. and C. Gaillardin (1997) Physiology and genetics of dimorphic fungus Yarrowia lipolytica. FEMS Microbiol. Rev. 19: 219–237.
Hassanshahian, M., H. Tebyanian, and S. Cappello (2012) Isolation and characterization of two crude oil-degrading yeast strains, Yarrowia lipolytica PG-20 and PG-32, from the Persian Gulf. Mar. Pollut. Bull. 64: 1386–1391.
Yano, Y., H. Oikawa, and M. Satomi (2008) Reduction of lipids in fish meal prepared from fish waste by a yeast Yarrowia lipolytica. Internat. J. Food Microbiol. 121: 302–307.
Madzak, C., C. Gaillardin, and J. M. Beckerich (2004) Heterologous protein expression and secretion in the non-conventional yeast Yarrowia lipolytica: A review. Biotechnol. J. 109: 63–81.
Juretzek, T., H.-J. Wang, J.-M. Nicaud, S. Mauersberger, and G. Barth, (2000) Comparison of promoters suitable for regulated overexpression of â-galactosidase in the alkane-utilizing yeast Yarrowia lipolytica. Biotechnol. Bioproc. Eng. 5: 320–326.
Guseva, M. A., E. Y. Epova, L. I. Kovalev, and A. B. Shevelev, (2010) The study of adaptation mechanisms of Yarrowia lipolytica yeast to alkaline conditions by means of proteomics. Appl. Biochem. Microbiol. 46: 307–312.
Vybornaya, T. V., T. V. Yuzbashev, T. I. Sobolevskaya, I. A. Laptev, E. Y. Yuzbasheva, and S. P. Sineoky (2011) Recombinant strain of the yeast Yarrowia lipolytica, producer of the lipase. Patent of Russian Federation 2,451,075.
Griffith, K. L. and Wolf, R. E. Jr. (1994) Measuring b-Galactosidase Activity in Bacteria: Cell Growth, Permeabilization, and Enzyme Assays in 96-Well Arrays. Biochem. Biophysic. Res. Communicat. 290: 397–402.
Miller, J. H. (1972) Experiments in Molecular Genetics. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA.
Blanchin-Rolland, S., R. R. Cordero-Otero, and C. Gallardin (1994) Two upstream activation sequences control the expression of the XPR2 gene in the yeast Yarrowia lipolytica. Mol. Cell Biol. 14: 327–338.
Varshavsky, A. (2001) Recent studies of the ubiquitin system and the N-end rule pathway. Harvey Lect. 96: 93–116.
Kobayashi, S., K. Hirakawa, H. Horiuchi, R. Fukuda, and A. Oht (2013) Phosphatidic acid and phosphoinositides facilitate liposome association of Yas3p and potentiate derepression of ARE1 (alkane-responsive element one)-mediated transcription control. Fungal. Genet. Biol. 61: 100–110.
Reyna-López, G. E., J. Simpson, and J. Ruiz-Herrera (1997) Differences in DNA methylation patterns are detectable during the dimorphic transition of fungi by amplification of restriction polymorphisms. Mol. Gen. Genet. 253: 703–710.
El-Maarri, O. (2003) Methods: DNA methylation. Adv. Exp. Med. Biol. 544: 197–204.
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Epova, E.Y., Balovneva, M.V., Isakova, E.P. et al. Expression system for Yarrowia lipolytica based on a promoter of the mitochondrial potential-dependent porin VDAC gene. Biotechnol Bioproc E 21, 408–413 (2016). https://doi.org/10.1007/s12257-016-0037-4
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DOI: https://doi.org/10.1007/s12257-016-0037-4