Isolation and Cleaning of Lactate Dehydrogenase from Pea (Pisum sativum L.) Roots by Hypoxia and the Study of Its Regulatory Properties

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The placement of pea plants (Pisum sativum L.) under flooding conditions led to an increase in lactate dehydrogenase (EC 1.1.1.27) activity in the roots. The enzyme was purified to an electrophoretic homogeneous state by a multistage purification method including ammonium sulfate fractionation, ion exchange chromatography on DEAE-Sephacel, and gel chromatography on Sephadex G-200. The degree of purification was 43.4, the yield was 2.5%, and the specific activity was 80.5 U/mg protein. Its physicochemical properties were studied: the molecular weight of the native lactate dehydrogenase molecule was 138 kDa. The molecular weight of the subunits was determined by PAGE by electrophoresis in the presence of DDS-Na. Its value was 34 kDa, which indicates that the enzyme is a homotetramer. The kinetic and regulatory properties of the enzyme and the values ​​of the Michaelis constants were established. he effect of the concentration of hydrogen ions and temperature on direct and reverse reactions catalyzed by it was obtained. It was determined that lactate dehydrogenase inhibited ATP.