Purification of NADH-Ferricyanide and NADH-Duroquinone Reductases from Maize (Zea mays L.) Root Plasma Membranes

Abstract

The plasma membrane of eukaryotic cells contains endogenous, integral electron transport activities. These activities include NAD(P)H: ferricyanide reductase (FCR) and NAD(P)H:duroquinone reductase (DQR). DQR and FCR exhibit unique properties with regard to specificity for pyridine nucleotides, degree of stimulation of activity by detergents, solubilization characteristics and degree of purification following affinity chromatography. Both activities have been effectively solubilized using Triton X-100 and partially resolved and purified using dye-ligand affmity chromatography on Cibacron blue 3G-A-agarose. Elution of Triton-solubilized activities bound to Cibacron blue 3G-A-agarose with 25 μM NADH resulted in a purified protein with FCR and DQR activities. Evidence suggests that DQR and FCR eluted from Cibacron blue 3G-A-agarose with 25μM NADH represent dual activities on the same protein, while a second enzyme with only FCR activity is eluted at higher concentrations of NADH.