Reactivation of Gx-NOX catalyst after inactivation induced by organic solvents. Gx-NOX (rhombus) was incubated with 60 vol% of dioxane in 10 mM sodium acetate buffer at pH 5 and 37°C for 18 h. The inactivated preparation was vacuum filtered and then reactivated by incubation in 10 mM sodium phosphate buffer at pH 7 and 65°C. The same protocol was repeated up to three times, the inactivation step was depicted on the plot as an arrow. Different samples of both inactivation and reactivation steps were withdrawn in order to analyze enzyme activity as described in Methods. The activity of different inactivation/reactivation cycles was normalized assigning 100% of relative activity to the initial activity of biocatalyst right before being inactivated for the first time.