Figure 2. Optimization of CnpB hydrolytic activity.
(a) SDS-PAGE analysis of NiNTA purified M. tuberculosis CnpB. (b-e) BNPP hydrolysis activity of CnpB. (b) 500 nM CnpB was incubated with 0.2 mM BNPP in the presence and absence of 0.1 mM indicated metal ions for 5 minutes at 37°C and absorbance at 405 nm was recorded. (c) Increasing concentrations of CnpB were incubated with 0.2 mM BNPP in the presence and absence of 0.1 mM MnCl2 at 37°C and absorbance at 405 nm was monitored. (d) 15 nM CnpB was incubated with 0.2 mM BNPP and increasing concentrations of MnCl2 at at 37°C and absorbance at 405 nM was monitored. (e) 15 nM CnpB was incubated with 0.2 mM BNPP and 0.1 mM MnCl2 in pH 5–9.5 reaction buffer at 37°C and absorbance at 405 nm was monitored. In all panels, data are presented as mean ± standard deviation of n=3 replicates.