Conserved pyridoxal protein that regulates Ile and Val metabolism.

Escherichia coli YggS is a member of the highly conserved uncharacterized protein family that binds pyridoxal 5'-phosphate (PLP). To assist with the functional assignment of the YggS family, in vivo and in vitro analyses were performed using a yggS-deficient E. coli strain (ΔyggS) and a purified form of YggS, respectively. In the stationary phase, the ΔyggS strain exhibited a completely different intracellular pool of amino acids and produced a significant amount of l-Val in the culture medium. The log-phase ΔyggS strain accumulated 2-ketobutyrate, its aminated compound 2-aminobutyrate, and, to a lesser extent, l-Val. It also exhibited a 1.3- to 2.6-fold increase in the levels of Ile and Val metabolic enzymes. The fact that similar phenotypes were induced in wild-type E. coli by the exogenous addition of 2-ketobutyrate and 2-aminobutyrate indicates that the 2 compounds contribute to the ΔyggS phenotypes. We showed that the initial cause of the keto acid imbalance was the reduced availability of coenzyme A (CoA); supplementation with pantothenate, which is a CoA precursor, fully reversed phenotypes conferred by the yggS mutation. The plasmid-borne expression of YggS and orthologs from Bacillus subtilis, Saccharomyces cerevisiae, and humans fully rescued the ΔyggS phenotypes. Expression of a mutant YggS lacking PLP-binding ability, however, did not reverse the ΔyggS phenotypes. These results demonstrate for the first time that YggS controls Ile and Val metabolism by modulating 2-ketobutyrate and CoA availability. Its function depends on PLP, and it is highly conserved in a wide range species, from bacteria to humans.

The implication of YggT of Escherichia coli in osmotic regulation.

An Escherichia coli mutant lacking three major K(+) uptake systems, Trk, Kup, and Kdp, did not grow under low K(+)and high Na(+) concentrations. The introduction of fkuA and of fkuB of a marine bacterium, Vibrio alginolyticus, has been reported to compensate for the growth defect by accelerating the rate of K(+) uptake (Nakamura, Katoh, Shimizu, Matsuba, and Unemoto, Biochim. Biophys. Acta, 1277, 201-208 (1996)). We investigated the function of unknown genes of E. coli, yggS and yggT, homologs of fkuA and fkuB respectively. E. coli TK2420 cells, which lack the three K(+) uptake systems, did not grow under high Na(+) or mannitol concentrations. The growth defect was compensated by the introduction of the yggT gene alone: yggS was not required. Here we found that YggT endowed E. coli cells with a tolerance for osmotic shock, and discuss a possible mechanism.