Investigation of electrophoretic properties and role of mitochondrial lipoxygenase of pea plants

Abstract

Using specific staining of proteins after electrophoretic separation of mitochondrial fraction proteins the investigations of lipoxygenase, its properties and activity in pea plant cells under various aerations were taken. Aims of the work were to prove the presence of lipoxygenase in mitochondria of seedlings, to determine electrophoretic mobility of enzyme and molecular mass, to study diverse aeration impact on properties and activity of this enzyme. As a study object two-week old hydroponically grown pea seedlings were used. Aerial part of seedlings was placed in different gas media for 3h. Cell compartments isolation was done by differential centrifugation method accompanied by fraction purity determination by appropriate markers. Presence of lipoxygenase was proved by specific staining of electrophoregrams with linoleate as a substrate, enzyme activity was estimated spectrophotometrically. Appliance of native PAGE in mitochondria of studied plants showed occurrence of only one molecular form of lipoxygenase. Increase of enzyme activity in mitochondria of seedlings under oxygen deficit was not linked to appearance of new enzyme isoforms or change of properties, including electrophoretic mobility. This may suggest that under hypoxic stress and high concentrations of CO₂ we observe an activity change of same molecular form of mitochondrial lipoxygenase. Herewith high concentrations of CO₂ increased also the activity of mitochondrial lipoxygenase in the studied plant that is evidenced by using specific inhibitors of this enzyme.

Supposed that enzymatic lipoxygenase path of hydroperoxide radicals in plant mitochondria makes significant contribution into processes of lipid peroxidation in plants under hypoxia but it is definitive only for rather small expositions (3-6 h).