Oxygen evolution by a reconstituted spinach chloroplast system in the presence ofl-glutamine and 2-oxoglutarate

Intact chloroplasts prepared from summer-grown spinach plants supported (aspartate plus 2-oxoglutarate)-dependent O₂ evolution but not (glutamine plus 2-oxoglutarate)-dependent O₂ evolution. The former activity, which was sensitive to amino oxyacetate, was attributed to transaminase activity and reduction of the resulting oxalo-acetate to malate using H₂O as eventual electron donor. A reconstituted chloroplast system which included chloroplast stroma, thylakoid membranes, ferredoxin and NADP(H) supported O₂ evolution in the presence ofl-glutamine and 2-oxoglutarate at rates of 15–22 μmol mg^-1 chlorophyll h^-1 although lower rates were obtained with material from winter-grown plants. Activity was not observed in the absence of ferredoxin and omission of NADP(H) decreased activity by 40%. The reaction was associated with the production of 0.49 mol O₂ mol^-1 2-oxoglutarate consumed and up to 0.46 mol O₂ mol^-1 glutamine supplied. The reaction, which was inhibited by azaserine but not by methionine sulphoximine or amino oxyacetate, was attributed to light-coupled glutamate synthase (EC 1.4.1.13) with H₂O serving as eventual electron donor. Activity was not affected significantly byl-malate. The reconstituted system also supported O₂ evolution in the presence of nitrite, oxaloacetate, (aspartate plus 2-oxoglutarate) and oxidised glutathione.