Human acylphosphatase cannot replace phosphoglycerate kinase in Saccharomyces cerevisiae

Area of Science:

• Biochemistry
• Molecular Biology
• Yeast Genetics

Background:

• Human acylphosphatase (h-AP) catalyzes 1,3-diphosphoglycerate hydrolysis.
• This reaction could bypass yeast phosphoglycerate kinase, potentially altering ATP production in glycolysis.
• Investigating h-AP's in vivo function in Saccharomyces cerevisiae is crucial for metabolic engineering.

Purpose of the Study:

• To determine if human acylphosphatase (h-AP) can functionally replace yeast phosphoglycerate kinase in Saccharomyces cerevisiae.
• To assess the impact of h-AP expression on glycolysis and ATP yield in S. cerevisiae.

Main Methods:

• A multi-copy plasmid with the h-AP gene under the TDH3 promoter was introduced into a pgk1 delta mutant of S. cerevisiae.
• Chemostat cultures were maintained at a dilution rate of 0.10 h(-1) using a glucose-ethanol mixture.
• Growth, glucose consumption, ethanol formation, and acylphosphatase activity were measured.

Main Results:

• The h-AP expressing strain showed high acylphosphatase activity in cell extracts.
• Switching to glucose as the sole carbon source led to complete growth arrest and cessation of glucose consumption.
• No ethanol was produced, indicating a non-functional glycolytic pathway.

Conclusions:

• Human acylphosphatase (h-AP) cannot substitute for yeast phosphoglycerate kinase in vivo.
• h-AP expression disrupts the glycolytic pathway in S. cerevisiae.
• h-AP is not a suitable tool for modifying ATP yield from glycolysis in yeast.