Rate-limiting factors in leaf photosynthesis. I. Carbon fluxes in the calvin cycle

Abstract

Rates of photosynthesis of spinach leaves were varied by varying light intensity and CO 2 concentration. Metabolism of the leaves was then arrested by freezing them in liquid nitrogen. Chloroplasts were isolated by a nonaqueous procedure. In the chloroplast fractions, levels of intermediates of the carbon reduction cycle were determined and considered in relation to the photosynthetic flux situation of the leaves at the time before freezing. During induction of photosynthesis, ribulose 1,5-bisphosphate levels increased in parallel with CO 2 fixation. In the steady state, a similar relation between ribulose 1,5-bisphosphate levels and CO 2 uptake was observed at light intensities between 0 and 50 W·m −2. A further increase in light intensity increased CO 2 fixation rates but not ribulose 1,5-bisphosphate levels. Increasing the CO 2 concentration resulted in increased CO 2 uptake, whereas ribulose 1,5-bisphosphate levels decreased. Even under CO 2 saturation, ribulose 1,5-bisphosphate levels were about 100 nmol/mg chlorophyll corresponding to about 3.5 mM ribulose 1,5-bisphosphate in the chloroplast stroma. This suggests that even under CO 2 saturation, ribulose-1,5-bisphosphate carboxylase limits photosynhetic CO 2 uptake. Mass action ratios calculated from measured metabolite levels demonstrated that the thermodynamic gradient required for the regeneration of ribulose 1,5-bisphosphate from hexosephosphate and triosephosphate increased considerably as photosynthetic flux increased. Similar calculations revealed that the enzymatic apparatus responsible for the reduction of 3-phosphoglycerate to dihydroxyacetone phosphate is not displaced much from equilibrium even under maximum rates of photosynthesis at saturating CO 2. The same is true for aldolase. Fructose-1,6-bisphosphatase also did not limit Calvin cycle turnover. Only at very low light intensities and during the first minutes of the induction period was the ratio of fructose 1,6-bisphosphate to fructose 6-phosphate high. This observation was more readily explained in terms of fructose 1,6-bisphosphate binding to ribulose-1,5-bisphosphate carboxylase than by a rate limitation imposed by insufficient activation of fructose-1,6-bisphosphatase.