Figure 3

Ammonium is released through the action of YbaS. (A) A schematic diagram of the ammonium measurement assay. Upon inoculation into LB medium supplemented with 0.4% glucose (to suppress AR1), E. coli was grown overnight to reach stationary phase. The cells were pelleted by centrifugation, washed with distilled water, pelleted again and resuspended in distilled water before mixing with 10 ml of citrate buffer, pH 2.5, supplemented with a specific amino acid. The ammonium concentration was continuously measured by a special meter. (B) Deletion of the ybaS gene resulted in no increase of ammonium concentration in the buffer. By contrast, ammonium concentration in the buffer increased steadily over time for WT or ΔyneH E. coli. 1 mM Gln was present in the buffer. (C) Ammonium concentration steadily increased only in the presence of Gln, but not Glu or Asn. (D) A higher concentration of Gln in the buffer gave rise to higher concentrations of ammonium. The results shown in panels (B-D) are the average of three independent experiments, with the SDs indicated.