FIGURE 3.

Substrate specificity of recombinant maize nucleotide pyrophosphatase/phosphodiesterase. Substrates (0.5 mM) were incubated with the purified enzyme in 100 mM McIlvaine buffer containing 20 mM MgCl₂ at 37°C for the period of 30 min, during which both the decrease in substrate concentration and increase in the product concentration were monitored by UFLC and the tangent was used to calculate the reaction rate. Graph depicts average of three technical replicates with standard deviation. The insert shows the saturation curve of FAD hydrolysis with maximal velocity on y-axis expressed in arbitrary units (a.u.), from which K_m of 32.7 ± 3.0 μM was calculated by GraFit software.