Changing Kinetic Properties of Fructose-1,6-bisphosphatase from Pea Chloroplasts during Photosynthetic Induction

Abstract

ABSTRACrAfter dark-light transitions, there is a delay in photosynthetic CO2 fixation by isolated pea chloroplasts in the range of some minutes. In order to assess the physiological significance of light modulation of enzyme activity in the control of induction, we made estimates of the kinetic parameters of fructose-1,6-bisphosphatase immediately upon release from pea chloroplasts in the dark and after illumination for various time periods. The Michaelis constant for fructose-1,6-bisphosphate decreased and maximal velocities increased during -induction. It seems likely that light activation of this enzyme is one of the factors contributing to the overcoming of the lag period in photosynthetic CO2 fixation.When isolated chloroplasts are exposed to dark-light transitions, there is a delay in photosynthetic CO2 fixation in the range of minutes. The length of the induction period depends on the incubation medium and on the condition of the chloroplasts (6,7,9,16,20). The question arises whether induction is mainly related to the concentration of stromal intermediates, to the activation of enzymes, or to both (5-7, 16, 17, 20, 21, 23). FruP2ase2 is thought to be one of the controlling enzymes of the reductive pentose phosphate pathway. The regulation of the enzyme activity in vivo has been suggested to be due to lightinduced changes in stromal Mg2+ concentration and pH (14), and to activation (presumably reductive covalent modification) mediated by light (1, 4). In experiments with purified spinach Fru-P2ase, it has been shown that DTT treatment does not change the maximal velocity of the enzyme, but does change the substrate dependence from sigmoidal to Michaelis Menten (5), so that the apparent affinity of the enzyme for both Fru-1,6-P2 and Mg2e is markedly increased. It therefore seemed of interest to determine the effect of light treatment on the kinetic parameters of the enzyme in intact chloroplasts.To assess the physiological significance of light modulation of Fru-P2ase during induction, we measured the kinetic parameters of the enzyme immediately after release from chloroplasts in the dark, at the end of the lag period, and twice during the maximal rate of C02-dependent 02 evolution. The results indicate that the light-effected properties of Fru-P2ase probably affect the increase of the C02-dependent 02 evolution rate during photosynthetic induction in vivo.