Purification and characterization of lipoxygenase from Entermorpha clathrata

摘要

The purification and characterization of lipoxygenase (LOX,EC 1.13.11.12) from the green algae Enteromorpha clathrata were studied. Two components marked LOX-1 and LOX-2 were purified and their molecular masses were estimated to be 102 and 79 ku by SDS-PAGE. Both LOX-1 and LOX-2 were stable over the pH wide range 6.0~10.0 and had the optimum pH of 10.0 and 8.6,at optimum temperature of 30 and 25℃,respectively. Substrate specificities of LOX-1 and LOX-2 were the greatest towards linoleic acid,followed by arachidonic acid and linolenic acid. The Michaelis constant values of LOX-1 and LOX-2 were 0.23 and 0.20 mmol/dm3 with the substrate of linoleic acid. The LOX activities were stimulated greatly by Ca2+ but inhibited by Hg2+ and the antioxidants such as BHA,BHT and TBHQ. The hydroperoxide products of LOX were analysed by HPLC with the substrate of methyl linoleate,and the results showed that LOX-1 formed mainly 9-hydroperoxides while LOX-2 formed both 9-and 13-hydroperoxides at a ratio of 24∶76.