Evaluation of Pyrophosphate‐Driven Proton Pumps in Saccharomyces cerevisiae under Stress Conditions

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Abstract

In Saccharomyces cerevisiae, pH homeostasis is reliant on ATP due to the use of proton-translocating ATPase (H+-ATPase) which constitutes a major drain within cellular ATP supply. Here, an exogenous proton-translocating pyrophosphatase (H+-PPase) from Arabidopsis thaliana, which uses inorganic pyrophosphate (PPi) rather than ATP, was evaluated for its effect on reducing the ATP burden. The H+-Ppase was localized to the vacuolar membrane or to the cell membrane, and their impact was studied under acetate stress at a low pH. Biosensors (pHluorin and mQueen-2m) were used to observe changes in intracellular pH (pHi) and ATP levels during growth on either glucose or xylose. A significant improvement of 35% in the growth rate at a pH of 3.7 and 6 g·L−1 acetic acid stress was observed in the vacuolar membrane H+-PPase strain compared to the parent strain. ATP levels were elevated in the same strain during anaerobic glucose and xylose fermentations. During anaerobic xylose fermentations, co-expression of pHluorin and a vacuolar membrane H+-PPase improved the growth characteristics by means of an improved growth rate (11.4%) and elongated logarithmic growth duration. Our study identified a potential method for improving productivity in the use of S. cerevisiae as a cell factory under the harsh conditions present in industry.
Original languageEnglish
Article number625
Number of pages23
JournalMicroorganisms
Volume12
Issue number3
DOIs
Publication statusPublished - 2024 Mar 20

Subject classification (UKÄ)

  • Microbiology
  • Industrial Biotechnology

Free keywords

  • Saccharomyces cerevisiae
  • pH homeostasis
  • ATP
  • proton-translocating ATPase
  • proton-translocating pyrophosphatase
  • pHluorin
  • mQueen-2m
  • acetic acid
  • xylose

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