The pyrG gene from Lactococcus lactis encodes CTP synthase (EC 188.8.131.52), an enzyme converting UTP to CTP. A series of strains were constructed with different levels of pyrG expression by insertion of synthetic constitutive promoters with different strengths in front of pyrG. These strains expressed pyrG levels in a range from 3 to 665% relative to the wild-type expression level. Decreasing the level of CTP synthase to 43% had no effect on the growth rate, showing that the capacity of CTP synthase in the cell is in excess in a wild-type strain. We then studied how pyrG expression affected the intracellular pool sizes of nucleotides and the correlation between pyrG expression and nucleotide pool sizes was quantified using metabolic control analysis in terms of inherent control coefficients. At the wild-type expression level, CTP synthase had full control of the CTP concentration with a concentration control coefficient close to one and a negative concentration control coefficient of -0.28 for the UTP concentration. Additionally, a concentration control coefficient of 0.49 was calculated for the dCTP concentration. Implications for the homeostasis of nucleotide pools are discussed.
|Journal||European Journal of Biochemistry|
|Publication status||Published - 2004|