The pathway of ammonia assimilation in Bacteroides fragilis.

Abstract

The enzymes in ammonia assimilation pathways have been examined in a strictly anaerobic bacterium Bacteroides fragilis. The activities of NADPH- and NADH-linked glutamate dehydrogenase and glutamine synthetase were demonstrated in extracts of cells, but very low activity of NADPH-dependent glutamate synthase and no activity of alanine dehydrogenase were demonstrated. Both activities of the glutamate dehydrogenase were not distinguished electrophoretically, indicating the presence of a dual pyridine nucleotide-specific enzyme. At low concentrations of ammonia in batch cultures and at low dilution rates of continuous flow cultures, higher activities of glutamate dehydrogenase were found in the cells. The values of Km of NADPH-linked glutamate dehydrogenase were 0.8mM, 0.15mM, and 7μM for ammonia, 2-oxoglutarate, and NADPH, respectively. Although glutamine synthetase activity was also higher in cultures with limited ammonia and at low dilution rates of continuous cultures, this enzyme may not be important for ammonia incorporation into amino acids, since cell growth was not affected by the addition to the culture of methionine sulfoximine, a glutamine synthetase inhibitor. This evidence suggests that ammonia assimilation is mainly carried out by the glutamate dehydrogenase in B. fragilis.