31P NMR magnetization-transfer measurements of flux between inorganic phosphate and adenosine 5'-triphosphate in yeast cells genetically modified to overproduce phosphoglycerate kinase.
31P NMR magnetization-transfer measurements were used to measure flux between inorganic phosphate and ATP in the reactions catalyzed by phosphoglycerate kinase and glyceraldehyde-3-phosphate dehydrogenase in anaerobic cells of the yeast Saccharomyces cerevisiae. Flux between ATP and Pi and glucose consumption and ethanol production were measured in cells expressing different levels of phosphoglycerate kinase activity. Overexpression of the enzyme was obtained by transforming the cells with a multicopy plasmid containing the phosphoglycerate kinase coding sequence and portions of the promoter element. Fluxes were also measured in cells in which the glyceraldehyde-3-phosphate dehydrogenase activity had been lowered by limited incubation with iodoacetate. These measurements showed that both enzymes have low flux control coefficients for glycolysis but that phosphoglycerate kinase has a relatively high flux control coefficient for the ATP----Pi exchange catalyzed by the two enzymes. The Pi----ATP exchange velocities observed in the cell were shown to be similar to those displayed by the isolated enzymes in vitro under conditions designed to mimic those in the cell with respect to the enzyme substrate concentrations.
Brindle
K M
KM
Department of Biochemistry, University of Oxford, England.
D000227:Adenine Nucleotides / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism*; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000378:metabolism; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D009243:NAD / Q000378:metabolism; D010710:Phosphates / Q000378:metabolism*; D010735:Phosphoglycerate Kinase / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*
D000227:Adenine Nucleotides; D010710:Phosphates; D009243:NAD; D000255:Adenosine Triphosphate; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D010735:Phosphoglycerate Kinase