Effect of Light on Chemical Modification of Chloroplast Ferredoxin-NADP Reductase

Chemical modification of spinach chloroplasts by phenylglyoxal and dansyl chloride resulted in inhibition of NADP photoreduction. The rate of inactivation was higher with both reagents when modification was carried out in the light with methylviologen or phenazine methosulfate present. Uncouplers prevent the effect of light. Electron transport from water to methylviologen was not affected by the modifiers. The presence of 10 millimolar NADP completely protected the membrane-bound reductase against inactivation by phenylglyoxal. With lower concentrations, protection was higher in the light than in the dark. The apparent dissociation constants of the enzyme-substrate complex for NADP were 0.9 and 0.1 millimolar for the dark and light inactivation, respectively. Inactivation of NADP photoreduction by dansyl chloride was completely prevented by ferredoxin, but only partially by nucleotides. The diaphorase activity was inhibited in chloroplasts modified by phenylglyoxal, but not when modified by dansyl chloride. The results suggest that energizing thylakoid membranes by light induces a conformational change in membrane-bound ferredoxin-NADP reductase, and that the reductase is an allotopic enzyme.