Exploring the potential of the glycerol-3-phosphate dehydrogenase 2 (GPD2) promoter for recombinant gene expression in Saccharomyces cerevisiae

Jan Dines Knudsen, Ted Johanson, Anna Eliasson Lantz, Magnus Carlquist

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Abstract

A control point for keeping redox homeostasis in Saccharomyces cerevisiae during fermentative growth is the dynamic regulation of transcription for the glycerol-3-phosphate dehydrogenase 2 (GPD2) gene. In this study, the possibility to steer the activity of the GPD2 promoter was investigated by placing it in strains with different ability to reoxidise NADH, and applying different environmental conditions. Flow cytometric analysis of reporter strains expressing green fluorescent protein (GFP) under the control of the GPD2 promoter was used to determine the promoter activity at the single-cell level. When placed in a gpd1Δgpd2Δ strain background, the GPD2 promoter displayed a 2-fold higher activity as compared to the strong constitutive glyceraldehyde-3-phosphate dehydrogenase (TDH3). In contrast, the GPD2 promoter was found to be inactive when cells were cultivated in continuous mode at a growth rate of 0.3 h-1 and in conditions with excess oxygen (i.e. with an aeration of 2.5 vvm, and a stirring of 800 rpm). In addition, a clear window of operation where the gpd1Δgpd2Δ strain can be grown with the same efficiency as wild type yeast was identified. In conclusion, the flow cytometry mapping revealed conditions where the GPD2 promoter was either completely inactive or hyperactive, which has implications for its implementation in future biotechnological applications such as for process control of heterologous gene expression.
Original languageEnglish
Journal Biotechnology Reports
Volume7
Pages (from-to)107-119
Number of pages13
DOIs
Publication statusPublished - 2015

Bibliographical note

This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

Keywords

  • Flow cytometry
  • Glycerol-deficient yeast
  • GPD deletion
  • GPD2 promoter
  • Inducible promoter
  • NADH/NAD+
  • Population heterogeneity
  • Recombinant protein production
  • Redox reporter

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