Effects of Protector and Hydroxyapatite Partial Purification on Stability of Lipoxygenase from Gray Mullet Gill

Three lipoxygenase (LOX, EC 1.13.11.12) isozymes, 5-, 12-, and 15-LOX, were found in the gill of cultured gray mullet ( Mugil cephalus ). The major LOX in mullet gill was the 12-isozyme followed by the 15-isozyme. Crude mullet gill extract (using 0.05 M potassium phosphate buffer, pH 7.0) catalyzed oxidation of arachidonic acid at an oxygen consumption rate of 30.6 ± 7.1 nmol min - 1 (mg of protein) -1 at optimal pH 7.0 and 25 °C. The half-life of LOX was 7−80 h at 4 °C depending on the postmortem time of the mullet. The mullet gill LOX was a very labile protein and lost activity during or after purification. Adding imidazole, histidine, glutathione, and trimethylamine increased the stability of LOX significantly. Optimal quantity of imidazole was 2.5 mmol/g of protein. Hydroxyapatite purification resulted in a 10.75-fold specific activity and maintained >80% activity at 4 °C for 3 days.