Simultaneous genomic overexpression of seven glycolytic enzymes in the yeast Saccharomyces cerevisiae |
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Authors: | Hauf Zimmermann Müller |
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Affiliation: | a Institut für Mikrobiologie und Genetik, Technische Universität Darmstadt, Schnittspahnstrasse 10, D-64287 Darmstadt, Germany b Scientific Research and Development GmbH, Oberurseler Strasse 43, D-61440 Oberursel, Germany c Stratagene Europe, Gebouw California, Hogehilweg 15, 1101 CB Amsterdam Zuidoost, The Netherlands |
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Abstract: | Fusions of the glycolytic genes TPI1, PGK1, ENO1, PYK1, PDC1, and ADH1 with the lacZ reporter gene of Escherichia coli and a lacZ fusion construct of a 390-bp fragment from the promoter of the HXT7 gene were assayed for β-galactosidase activity. The glycolytic promoters were induced after addition of glucose to ethanol-grown cells, whereas the HXT7 promoter fragment showed a constitutive β-galactosidase expression on both carbon sources. The genes coding for the seven enzymes of lower glycolysis Tdh, Pgk, Gpm, Eno, Pyk, Pdc, and Adh were simultaneously put under the control of the same strong promoter, a truncated HXT7 promoter that is constitutively active on ethanol as well as on glucose medium. Genomic expression of the glycolytic genes under the control of this promoter, resulted in an at least 2-fold overexpression. The gene MSG5 was isolated, coding for a protein phosphatase normally involved in cell cycle regulation, as a factor that possibly influences the expression of the HXT7 gene. However, overexpression of MSG5 had no effect on the expression of the HXT7/lacZ fusion, whereas a deletion of this gene resulted in a decreased expression of β-galactosidase. |
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Keywords: | Saccharomyces cerevisiae Glycolysis Overexpression |
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