Purification and properties of lipoxygenase from opium poppy seedlings(Papaver somniferum L.)

Authors : İvana HOLKOVÁ, Frantisek BILKA, Drahomíra RAUOVÁ, Lydia BEZÁKOVÁ

Pages : 772-780

View : 16 | Download : 10

Publication Date : 2016-12-01

Article Type : Research Paper

Abstract :Lipoxygenase insert ignore into journalissuearticles values(linoleate:oxygen oxidoreductase; EC 1.13.11.12; LOX); is a key enzyme in the signaling pathway leading to the production of biologically active compounds involved in plant growth development and stress responses. In this study the novel LOX enzyme from opium poppy insert ignore into journalissuearticles values(Papaver somniferum L.); seedlings was purified to electrophoretic homogeneity and characterized. LOX activity during the germination of opium poppy seeds was analyzed, and the highest LOX activity was determined on the fourth day of germination. Opium poppy LOX was purified from 4-day-old poppy seedlings after ammonium sulfate precipitation followed by hydrophobic chromatography insert ignore into journalissuearticles values(Phenyl-Sepharose CL-4B);, ion exchange chromatography insert ignore into journalissuearticles values(Q-Sepharose);, and affinity chromatography insert ignore into journalissuearticles values(linoleyl-aminopropyl agarose); for the first time. The relative molecular weight of purified LOX was estimated to be 78 kDa by immunoblotting. The highest enzyme activity occurred at pH 6.5 and 0.75 mM calcium ion concentration. LOX showed preferential activity towards linoleic acid followed by linolenic acid as a substrate. To investigate the positional specificity of the LOX reaction, purified LOX was incubated with linoleic acid, and the products were analyzed by high-performance liquid chromatography. Opium poppy LOX converted linoleic acid to 13-hydroperoxy-insert ignore into journalissuearticles values(9Z,11E);-octadecadienoic acid insert ignore into journalissuearticles values(63.5%); and, to a lesser extent, 9-hydroperoxy-insert ignore into journalissuearticles values(10E,12Z);-octadecadienoic acid insert ignore into journalissuearticles values(36.5%);.
Keywords : Lipoxygenase, enzyme purification, biochemical characterization, positional specificity, Papaver somniferum