Regulation of PRPP and nucleoside tri and tetraphosphate pools in Escherichia coli under conditions of nitrogen starvation

Abstract

The ribonucleoside triphosphate, deoxyribonucleoside triphosphate, guanosine 3''-diphosphate 5''-diphosphate (ppGpp), and 5-phosphoribosyl 1-pyrophosphate (PRPP) pools in E. coli B were determined by TLC during changing conditions to ammonium starvation. The intracellular concentrations of all nucleotides changed in a well-defined order several minutes before any observed change in the optical density of the culture. The levels of purine nucleoside triphosphates (ATP, dATP, and GTP) decreased, followed by a sequential decrease in cytidine nucleotides (CTP, dCTP) and uridine nucleotides (UTP and dTTP). The deoxyribonucleotides thus behaved as the ribonucleotides. The levels of ppGpp increased 11-fold after the decrease in uridine nucleotides, when the accumulation of stable RNA stopped. The level of the nucleotide pool did not stabilize until 30 min after the change in optical density. The pool of dGTP dropped concomitantly with the pool of CTP. The nucleotide precursor PRPP exhibited a transient increase, with maximum value of 4 times the exponential levels at the onset of starvation. Apparently, the cell adjusts early to starvation by reducing the phosphorylating activity or the nucleotide biosynthetic activity. As in other downshift systems, the accumulation of stable RNA stopped before the break in optical density and before the stop in protein accumulation. Cell divisions were quite insensitive to control mechanisms operating on RNA and protein accumulation under ammonium starvation, since the cells continued to divide for 21 min without any net accumulation of RNA.