Orthophosphate Control of Glucose-6-Phosphate Dehydrogenase Light Modulation in Relation to the Induction Phase of Chloroplast Photosynthesis

Recently, an increase in work concerned with the organization of enzymes in a semi-or completely immobilized form within the living cell has appeared (17). Three ofthe enzymes ofCO2 fixation, ribulose-P2 carboxylase (18), ribulose-5-P kinase (1 1), and aldolase (5), are at least partially associated with the thylakoid membrane in spinach or pea chloroplasts. In spinach chloroplasts, G6PD4 (EC 1.1.1.49) (24) and NADP-linked malic dehydrogenase (23) also are partitioned between soluble and membrane fractions. The purpose of the present experiments was to examine the association of G6PD with membranes in the pea leaf chloroplast.We have examined the effect of high pH, Pi, other salts, various metabolites, and light on the association of G6PD with the particulate fraction of the chloroplast. We conclude that the membranebound form of the enzyme is a peripheral protein. Light affects the partitioning ofthe enzyme between the membrane and stromal portions of the chloroplast. This effect may be related to the light inactivation of the enzyme (1). MATERIALS AND METHODSChloroplast Preparation. Pea (Pisum sativum L., var. Little Marvel) plants were grown in vermiculite in a greenhouse. Chloroplasts were prepared from young (9-to 11-day-old) shoots as described by Cockbum et al. (9), but in the absence of sodium isoascorbate, and washed once in 0.33 M sorbitol, 50 mM Hepes

mentioning

confidence: 79%

Light-Induced Release of Bound Glucose-6-phosphate Dehydrogenase to the Stroma in Pea Chloroplasts

Ben-Bassat¹,

Anderson²

1981

“…Chloroplasts from 'rapidly' fractionated oat protoplasts had 3 mm Pi in the experiments of Hampp et al (10). Because of the Pi sensitivity of starch synthesis and enzyme activation (12,13), I believe that the Pi concentration inside the chloroplast can often be as low as 1 (22).…”

Section: Discussionmentioning

confidence: 99%

O₂-Insensitive Photosynthesis in C₃ Plants

Sharkey¹

1985

249

105

There have been a number of reports of no stimulation of photosynthesis and sometimes even an inhibition upon switching from air levels of 02 to low 02 pressure (29,31). This unusual behavior is sensitive to temperature (5, 14, 15), CO2 pressure, and light (18). In this paper, I show that as light is varied the inhibition of photosynthesis by 02 is nearly constant over a range of CO2 assimilation rates until the rate of CO2 assimilation approaches a maximum, whereupon the 02 inhibition becomes at first less severe, and then can even reverse. This 02-insensitive limitation to photosynthesis also occurs in water-stressed plants. The postillumination burst of CO2 was used to demonstrate that the lack of 02 sensitivity was not the result of reduced photorespiration. Since the 02-insensitive limitation of photosynthesis appears to set an upper limit on the rate of photosynthetic CO2 assimilation, it was studied in a weedy species with a high rate of assimilation (X. strumarium) and a crop species that has a maximum assimilation rate of less than half that of Xanthium (Phaseolus vulgaris). No differences in the nature of 02 insensitivity were noted, and results from both species are presented. Photosynthesis in plants having the C3 pathway of photosynthesis is known to be inhibited by air levels of 02 pressure when the partial pressure of CO2 is below 1000 ,bar. Present

“…Activation of reductive pentose phosphate cycle enzymes occurs at a comparable rate in this system (16). Huber (13) has suggested that glucose 6-P dehydrogenase activity might contribute to the buildup of intermediates during induction. However, in experiments which are comparable to the experiments reported here, we measured metabolite levels and found that glycerate 3-P, triose-P, hexose 1,6-bisP, sedoheptulose 1,7-bisP and ribulose 1,5-bisP levels remained low and apparently constant during the first 2 min of illumination (16).…”

mentioning

confidence: 99%

Changing Activity of Glucose-6-Phosphate Dehydrogenase from Pea Chloroplasts during Photosynthetic Induction

Yuan

Anderson²

1987

Light inactivation of glucose 6-phosphate dehydrogenase is rapid and occurs before photosynthetic 02 evolution is measureable in intact chloroplasts. Likewise, dark activation is rapid. The major light induced change in the kinetic parameters of glucose 6-phosphate dehydrogenase is in maximal velocity.The first enzyme of the oxidative pentose phosphate pathway, glucose 6-P dehydrogenase (EC 1.1.1.49), is inactivated when pea chloroplasts are irradiated (3). This light inactivation probably prevents operation of a futile cycle involving glucose 6-P, NADPH, and oxidative and reductive pentose phosphate pathway enzymes. We have now examined the kinetics of light inactivation of glucose 6-P dehydrogenase in intact chloroplasts during photosynthetic induction and the kinetic parameters of the active (dark) and less active (light) forms of the enzyme.Fickenscher and Scheibe (9) have partially purified cytosolic glucose 6-P dehydrogenase from peas. Unlike the purified chloroplastic enzyme (20), the cytosolic enzyme was not DTT or DTT/thioredoxin sensitive. On the basis ofthis lack of sensitivity to DTT and the results of antibody experiments with extracts from leaves which had been illuminated, Fickenscher and Scheibe concluded that the cytosolic enzyme was not light inactivated. But in their antibody experiments the error in activity determinations for the cytosolic enzyme from darkened and illuminated leaves was 18 and 23%, respectively (Fig. 3 in Ref. 9). Since errors are additive, the error in the determination of light inactivation was then the same as the expected change in activity if the cytosolic enzyme were 40% inactivated by light. Because previous work in this laboratory (4-6) indicated that the cytosolic enzyme was light and DTT inactivated, we have reinvestigated the DTT-dependent inactivation of the enzyme in whole leaf extracts. MATERIALS AND METHODSPhotosynthetic 02 Evolution and Determination of Kinetic Parameters. Chloroplasts were isolated from pea (Pisum sativum L. var Little Marvel) seedlings and photosynthetic 02 evolution assayed as described by Marques and Anderson (15) except that the chloroplast concentration was increased. Where activity with time of illumination was followed, the chloroplast content in the