Activity of CYP152BSβ at different H2O2 concentrations using the non-natural substrates ABTS (a,b) and guaiacol (c,d).
Activity assays were performed at room temperature (approx. 22°C) using heptanoic acid as decoy molecule. Product conversion was measured photometrically at 405 nm (ABTS) or 470 nm (guaiacol) and product formation was calculated using Lambert-Beer law (εABTS 36.8 mmol l-1 cm-1; εguaiacol 26.6 mmol l-1 cm-1). Specific activities were calculated based on the initial reaction velocities and given in mmol l-1 min-1 mg-1. Reactions were performed by adding up to 10 mM H2O2. For the conversion of ABTS, specific activity peaked at 1 mM, for guaiacol it peaked at 2.5 mM ((b,d) show blow-ups of the activity at low H2O2 concentrations, the dashed lines indicate the H2O2-dependent increase in specific activity). Means and standard deviations represent three independent experiments.