Fig. 5.

Abundance of pupylated proteins depends on the nitrogen source. (A) M. tuberculosis contains a lower abundance of pupylated proteins, and of a model PPS substrate, when grown in PB-nitrate compared with PB-Asn. Pupylated proteins were detected by immunoblot (IB) using a monoclonal antibody that recognizes M. tuberculosis Pup. The same immunoblot membranes were used to detect inositol-3-phosphate synthase (Ino1) and PrcB. The relative pupylome abundance between growth conditions (Bottom) was normalized by PrcB abundance and was statistically significant (Welch’s t test, P < 0.05). (B) Pup-Zur-His₆ levels are reduced in WT M. tuberculosis grown in PB-nitrate compared with PB-Asn. The normalized Pup-Zur-His₆ intensity in the second lane relative to the first lane is 0.62. PrcB and Mpa were detected on the same membrane, while PrcA was detected using a membrane separately prepared with the same lysates.