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 degrees C. The half-life of LOX was 7-80 h at 4 degrees 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 degrees C for 3 days.