KINETIC AND ELECTROPHORETIC CHARACTERIZATION OF NADP DEPENDENT DEHYDROGENASES FROM ROOT TISSUES OF NORWAY SPRUCE [PICEA-ABIES (L) KARST] EMPLOYING A RAPID ONE-STEP EXTRACTION PROCEDURE

The NADPH generating enzymes glucose-6-phosphate dehydrogenase (EC 1.1 .1.49), 6-phosphogluconate dehydrogepase (EC 1.1.1.44), and isocitrate -dehydrogenase (NADP dependent; EC 1.1.1.42) have been characterized i n spruce [Picea abies (L.) Karst.] roots. Interference from inherent p henolic compounds was minimized by complexation with borate and insolu ble polyvinylpyrrolidone in the presence of 2-mercaptoethanol and NADP . A further addition of protective substances had no (bovine serum alb umin) or even an inhibitory effect (ascorbate) on the enzyme activitie s. The enzymes were shown to be strictly NADP specific. The optimal pH -values and the apparent Michaelis constants from spruce roots are in good agreement with data from different photosynthetic organisms and g ametophytic tissues of conifers. Native electrophoresis and subsequent activity staining showed the same banding patterns for enzymes both f rom root and needle tissues. In addition, the applicability of a highl y sensitive dot-blot assay for the accurate quantification of the extr acted protein is shown.