Figure 3
Activities of the Mur enzymes (MurA-MurF) at various concentrations.
(A) Activity of MurA. The reaction contained 0.2 mM UDP-GlcNAc, 0.2 mM PEP and MurA enzyme at various concentrations. The X-axis represents the concentration of MurA enzyme (nM) and the Y-axis represents net Pi (nmol Pi/min). (B) Activity of MurB. The MurA reaction after a 30 min of incubation time was supplemented with 10 mM KCl, 0.2 mM NADPH and MurB enzyme at various concentrations in a total volume of 100 μl. The absorbance at 340 nm was monitored over 5 min to measure the rate of NADPH oxidation. The X-axis represents incubation time (seconds) and the Y-axis represents absorbance at 340 nm. (C–F) Activity of Mur ligases. The MurB reaction after a 5 min of incubation time was supplemented with 10 mM MgCl2, 1 mM L-Alanine, 2 mM ATP and MurC enzyme at various concentrations. The reaction was incubated at 37 °C for 60 minutes. Likewise for MurD assay, MurC reaction was supplemented with 1 mM D-Glutamate and MurD enzyme, for MurE assay MurD reaction was supplemented with 1 mM meso-Diaminopimelic acid and MurE enzyme and for MurF assay, MurE reaction was supplemented with 1 mM D-Alanine-D-Alanine and MurF enzyme. (C) MurC, (D) MurD (E) MurE (F) MurF. The X-axis represents a Mur ligases at various concentrations (nM) and the Y-axis represents net Pi (nmol Pi/min). On the bar/graph, “*” indicates the concentration of enzyme that was selected to be used subsequently in the respective assays. Data depicted is the mean ± S.E. values obtained from three independent experiments.
