Figure 5.
Effect of PII protein on NadE2 activity depends on the PII variant and is conserved in different bacteria. A, the activity of AbNadE2 was measured in the presence of MgCl2 5 mm, 1 mm ATP, 2.5 mm NAD+, and the indicated PII protein (GlnZ, GlnZΔloopT, GlnB, and GlnZ-UMP3) at 2 μm concentration (trimer), 1.5 mm 2-OG was added when indicated. Treatment without PII was carried with 2 μm of BSA. Activity measured following PPI production. An asterisk indicates the statistically significant difference compared with GlnZ (t test, p < 0.01), n = 3. B and C, NadE2 activity data were plotted as relative percent activity considering the reaction without NAD+ as 100% activity. All reactions were carried out in the presence of 100 nm HsNadE2 (B) or ScNadE2 (C), 4 mm ATP, 2 mm NaAD, and 4 mm l-glutamine. When indicated, the corresponding PII protein was added at 2 μm (trimer). Curves without PII were carried out using 2 μm BSA. Activity measured following PPI with EnzChek Pyrophosphate Assay Kit. An asterisk indicates a statistically significant difference compared (t test, p < 0.01); n = 3.