Glyceraldehyde-3-phosphate dehydrogenase has no control over glycolytic flux in Lactococcus lactis MG1363.
Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) has previously been suggested to have almost absolute control over the glycolytic flux in Lactococcus lactis (B. Poolman, B. Bosman, J. Kiers, and W. N. Konings, J. Bacteriol. 169:5887-5890, 1987). Those studies were based on inhibitor titrations with iodoacetate, which specifically inhibits GAPDH, and the data suggested that it should be possible to increase the glycolytic flux by overproducing GAPDH activity. To test this hypothesis, we constructed a series of mutants with GAPDH activities from 14 to 210% of that of the reference strain MG1363. We found that the glycolytic flux was unchanged in the mutants overproducing GAPDH. Also, a decrease in the GAPDH activity had very little effect on the growth rate and the glycolytic flux until 25% activity was reached. Below this activity level, the glycolytic flux decreased proportionally with decreasing GAPDH activity. These data show that GAPDH activity has no control over the glycolytic flux (flux control coefficient = 0.0) at the wild-type enzyme level and that the enzyme is present in excess capacity by a factor of 3 to 4. The early experiments by Poolman and coworkers were performed with cells resuspended in buffer, i.e., nongrowing cells, and we therefore analyzed the control by GAPDH under similar conditions. We found that the glycolytic flux in resting cells was even more insensitive to changes in the GAPDH activity; in this case GAPDH was also present in a large excess and had no control over the glycolytic flux.
Solem
Christian
C
Section of Molecular Microbiology, BioCentrum-DTU, Technical University of Denmark, DK-2800 Kgs. Lyngby, Denmark.
Koebmann
Brian J
BJ
Jensen
Peter R
PR
D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D002455:Cell Division / Q000235:genetics; D024601:Glyceraldehyde 3-Phosphate Dehydrogenase (NADP+) / Q000235:genetics / Q000378:metabolism*; D006019:Glycolysis; D013294:Lactococcus lactis / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation
D001426:Bacterial Proteins; D024601:Glyceraldehyde 3-Phosphate Dehydrogenase (NADP+); D000251:Adenosine Triphosphatases