Photoinhibition of CO(2)-Dependent O(2) Evolution by Intact Chloroplasts Isolated from Spinach Leaves

Intact spinach (Spinacia oleracea L.) chloroplasts, when pre-illuminated at 4 millimoles quanta per square meter per second for 8 minutes in a CO(2)-free buffer at 21% O(2), showed a decrease (30-70%) in CO(2)-dependent O(2) evolution and (14)CO(2) uptake. This photoinhibition was observed only when the O(2) concentration and the quantum fluence rate were higher than 4% and 1 millimole per square meter per second, respectively. There was only a small decrease in the extent of photoinhibition when the CO(2) concentration was increased from 0 to 25 micromolar during the treatment, but photoinhibition was abolished when the CO(2) concentration was increased to 30 micromolar. Addition of small quantities of P-glycerate (40-200 micromolar) or glycerate (160 micromolar) was found to prevent photoinhibition. Other intermediates of the Calvin cycle (fructose-6-P, fructose-1,6-P, ribose-5-P, ribulose-5-P) also prevented photoinhibition to various extents. Oxaloacetate was not effective in preventing photoinhibition in these chloroplasts. The amount of O(2) evolved during treatments with 3-P-glycerate or glycerate was no more than 65% of that measured in the presence of low CO(2) concentrations (9-12 micromolar) which did not prevent photoinhibition. In all cases, the extent to which photoinhibition was prevented by these metabolites was not correlated to the amount of O(2) evolved during the photoinhibitory treatment. It is concluded that in these chloroplasts the prevention of the O(2)-dependent photoinhibition of light saturated CO(2) fixation capacity is not linked to the dissipation of excitation energy via the photosynthetic electron transport nor to ATP utilization. The requirement of O(2) for photoinhibition of CO(2) fixation capacity in isolated chloroplasts may be explained by an effect of O(2) in allowing metabolic depletion of Calvin cycle intermediates.