Lysine decarboxylase activity assay

The activity of LDC was determined by measuring residual concentration of _L-lysine using lysine oxidase and peroxidase. After LDC reaction, lysine oxidase converts remaining lysine into 6-amino-2-oxohexanoate, NH₃, and H₂O₂ and then the hydrogen peroxide is reduced by peroxidase with 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS). The oxidized ABTS is detected by spectrophotometric method in absorbance at 412 nm. The assay was performed at 30 °C in a total volume 200 μl, containing 100 mM potassium phosphate, pH 6.0, 0.1 M _L-lysine, 0.2 mM pyridoxal-5-phosphate, and 25 μg of purified enzymes. The reaction was stopped by heating the reaction mixture at 100 °C for 5 min. After centrifugation at 13,500 × g for 1 min, 2X reaction solution that contains 0.1 unit ml⁻¹ lysine oxidase and 1 unit ml⁻¹ peroxidase in potassium phosphate buffer is added to the reaction mixture.