Abstract
In order to identify subunit composition and substrate specificity of the plastid-type Complex I (proton-translocating NAD(P)H dehydrogenase), we isolated chromoplasts from red fruits of paprika. Membrane proteins were solubilized from thylakoid membranes with sucrose monolaurate, and NAD(P)H dehydrogenases were separated by anion-exchange HPLC. Peak 1 exhibited the high Km value (0.97 mM) for both NADH and NADPH, wheras Peak 2 with a clear pH optimum at 6.5 showed a higher affinity for NADH (Km, 0.28 mM) than NADPH. FNR was immunochemically detected between Peaks 1 and 2. An apparent molecular weight of the Peak 2 enzyme was estimated to be about 0.4 MDa by native polyacrylamide gel electrophoresis and 0.3 MDa by gel filtration, thus Peak 2 is likely to be the plastid-type Complex I, which participates in both cyclic photophosphorylation and chlororespiration. Purification of these enzymes and characterization of their substrate specificity are in progress.
