Fig. 1.
Characterization of WT PodA30–162 and PodA10. (A) WT PodA30–162 specific activity with pH, salt, and viscosity sweeps. WT PodA30–162 (3 µM) was incubated with PYO (100 µM), and activity was monitored by measuring absorbance of PYO (690 nm) over time. (B) PYO (50 µM) demethylation activity of PodA variants (3 µM) was determined using a spectrophotometric assay described in Materials and Methods. (C) Michaelis–Menten kinetics of PodA8 and PodA10 (3 µM of protein per reaction) were found by plotting Vmax values against [PYO], as indicated on the x axis. (D) PodA10 specific activity under varying conditions. PodA10 (3 µM) was incubated with PYO (100 µM), and activity was monitored by measuring absorbance of PYO over time. Activity was analyzed by changing pH, viscosity, or salt concentrations. Error bars in each panel represent SD of triplicates.