*3.6. Enzyme Activity Assay*

The enzyme activity of CamPhoD was determined at 37 ◦C for 30 min with the use of 2 mM p-nitrophenyl phosphate (*p*-NPP), thymidine-5 -monophosphate-p-nitrophenylester (5 -pNP-TMP) or bis-p-nitrophenyl phosphate (Bis-*p*-NPP) as chromogenic substrates in 25 mM Tris-HCl buffer, pH 9.0, 2 mM CoCl2, 2 mM FeCl3. The volume of the reaction mixture was 0.5 mL. The reaction was stopped by adding 1 mL of cooled 0.5 M NaOH to the reaction mixture. The absorption of the formed p-nitrophenol was measured at an optical density of 400 nm. The amount of enzyme required for the conversion of 1 μM *p*-nitrophenyl from a substrate over 1 min was taken as a unit of activity. The specific activity is given in units calculated per 1 mg of protein. The protein concentration was determined using Bradford's method [60].
