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Light-dependent reduction of dehydroascorbate and uptake of exogenous ascorbate by spinach chloroplasts

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Abstract

A reconstituted spinach chloroplast system containing thylakoids, stroma and 0.1 mM NADPH supported O2 evolution in the presence of oxidised glutathione (GSSG). The properties of the reaction were consistent with light-coupled GSSG-reductase activity involving H2O as eventual electron donor. The reconstituted system also supported dehydroascorbate-dependent O2 evolution in the presence of 0.6 mM reduced glutathione (GSH) and 0.1 mM NADPH with the concomitant production of ascorbate. The GSSG could replace GSH in which case the production of GSH preceded the accumulation of ascorbate. The data are consistent with the light-dependent reduction of dehydroascorbate using H2O as eventual electron donor via the sequence H2O→NADP→GSSG→dehydroascorbate. Approximately 30% of the GSH-dehydrogenase activity of spinach leaf protoplasts is localised in chloroplasts: this could not be attributed to contamination of chloroplasts by activity from the extrachloroplast compartment. Washed intact chloroplasts supported the uptake of ascorbate but the uptake mechanism had a very low affinity for ascorbate (Km approximately 20 mM). The rate of uptake of ascorbate was less than the rate of light-dependent reduction of dehydroascorbate and too slow to account for the rate of H2O2 reduction by washed intact chloroplasts.

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Abbreviations

Chl:

chlorophyll

GSH:

reduced glutathione

GSSG:

oxidised glutathione. In the text, glutathione refers collectively to GSH plus GSSG

References

  • Allen, J.F. (1977a) Oxygen — A physiological electron acceptor in photosynthesis. Curr. Adv. Plant Sci. 29, 459–469

    Google Scholar 

  • Allen, J.F. (1977b) Effects of washing and osmotic shock on catalase activity of intact chloroplast preparations. FEBS Lett. 84, 221–224

    Google Scholar 

  • Bradbeer, J.W. (1969) The activities of the photosynthetic carbon cycle enzymes of greening bean leaves. New Phytol. 68, 233–245

    Google Scholar 

  • Edwards, G.E., Robinson, S.P., Tyler, N.J.C., Walker, D.A. (1978) Photosynthesis by isolated protoplasts, protoplast extracts and chloroplasts of wheat. Influence of orthophosphate, pyrophosphate and adenylates. Plant Physiol. 62, 313–319

    Google Scholar 

  • Foyer, C.H., Halliwell, B. (1976) The presence of glutathione and glutathione reductase in chloroplasts: a proposed role in ascorbic acid metabolism. Planta 133, 21–25

    Google Scholar 

  • Foyer, C.H., Halliwell, B. (1977) Purification and properties of dehydroascorbate reductase from spinach leaves. Phytochemistry 16, 1347–1350

    Google Scholar 

  • Foyer, C.H., Rowell, J., Walker, D.A. (1983) Measurement of the ascorbate content of spinach leaf protoplasts and chloroplasts during illumination. Planta 157, 239–244

    Google Scholar 

  • Fridovich, I. (1975) Superoxide dismutases. Annu. Rev. Biochem. 44, 147–159

    Google Scholar 

  • Halliwell, B. (1974) Superoxide dismutase, catalase and glutathione peroxidase: solutions to the problems of living with O2. New Phytol. 73, 1075–1086

    Google Scholar 

  • Halliwell, B., Foyer, C.H. (1978) Properties and physiological function of a glutathione reductase purified from spinach leaves by affinity chromatography. Planta 139, 9–17

    Google Scholar 

  • Heldt, H.W. (1980) Measurement of metabolite movement across the envelope and of the pH in the stroma and the thylakoid space in intact chloroplasts. Methods Enzymol. 69, 604–613

    Google Scholar 

  • Jablonski, P.P. (1981) Reduction of hydrogen peroxide and selenite in illuminated pea chloroplasts. Ph.D. thesis, La Trobe University, Melbourne, Australia

  • Jablonski, P.P., Anderson, J.W. (1978) Light-dependent reduction of oxidized glutathione by ruptured chloroplasts. Plant Physiol. 61, 221–225

    Google Scholar 

  • Jablonski, P.P., Anderson, J.W. (1981) Light-dependent reduction of dehydroascorbate by ruptured pea chloroplasts. Plant Physiol 67, 1239–1244

    Google Scholar 

  • Jablonski, P.P., Anderson, J.W. (1982) Light-dependent reduction of hydrogen peroxide by ruptured pea chloroplasts. Plant Physiol 69, 1407–1413

    Google Scholar 

  • Kaiser, M.W. (1976) The effect of hydrogen peroxide on CO2 fixation of isolated chloroplasts. Biochim. Biophys. Acta 440, 476–482

    Google Scholar 

  • Law, M.Y., Halliwell, B. (1983) Glutathione and ascorbic acid in spinach (Spinacia oleracea) chloroplasts. The effect of hydrogen peroxide and of paraquat. Biochem. J. 210 (in press)

  • Lilley, R. McC., Fitzgerald, M.P., Rienits, K.G., Walker, D.A. (1975) Criteria of intactness and photosynthetic activity of spinach chloroplast preparations. New Phytol. 75, 1–10

    Google Scholar 

  • Lilley, R. McC., Walker, D.A. (1979) Studies with the reconstituted chloroplast system. In: Encyclopedia of plant physiology, N.S., vol. 6: Photosynthesis II. Photosynthetic carbon metabolism and related processes, pp. 41–52, Gibbs, M., Latzko, E., eds. Springer, Berlin Heidelberg New York

    Google Scholar 

  • Mapson, L.W., Isherwood, F.A. (1963) Glutathione reductase from germinated peas. Biochem. J. 86, 173–191

    Google Scholar 

  • Mehler, A.H. (1951) Studies on reactions of illuminated chloroplasts I. Mechanism of the reduction of oxygen and other Hill reagents. Arch. Biochim. Biophys. 33, 65–67

    Google Scholar 

  • Nakano, Y., Asada, K. (1980) Spinach chloroplasts scavenge hydrogen peroxide on illumination. Plant Cell Physiol 21, 1295–1307

    Google Scholar 

  • Nakano, Y., Asada, K. (1981) Hydrogen peroxide is scavenged by ascorbate-specific peroxidase in spinach chloroplasts Plant Cell Physiol. 22, 867–880

    Google Scholar 

  • Owens, C.W.I., Belcher, R.V. (1965) A colorimetric micro method for the determination of glutathione. Biochem. J. 94, 705–711

    Google Scholar 

  • Robinson, S.P., Walker, D.A. (1979) Rapid separation of the chloroplast and cytoplasmic fractions from intact leaf protoplasts. Arch. Biochem. Biophys. 196, 319–333

    Google Scholar 

  • Schaedle, M., Bassham, J.A. (1977) Chloroplast glutathione reductase. Plant Physiol. 59, 1011–1012

    Google Scholar 

  • Slabas, A.R., Walker, D.A. (1976) Inhibition of spinach phosphoribulokinase by DL-glyceraldehyde. Biochem. J. 153, 613–619

    Google Scholar 

  • Smith, J., Shrift, A. (1979) Phylogenetic distribution of glutathione peroxidase. Comp. Biochem. Physiol. B 63, 39–44

    Google Scholar 

  • Stadtman, T.C. (1979) Some selenium-dependent biochemical processes. Adv. Enzymol. 48, 1–28

    Google Scholar 

  • Tagawa, K., Arnon, D.I. (1962) Ferredoxins as electron carriers in photosynthesis and in the biological production and consumption of hydrogen gas. Nature (London) 195, 537–543

    Google Scholar 

  • Walker, D.A. (1980) Preparation of higher plant chloroplasts. Methods Enzymol. 69, 94–104

    Google Scholar 

  • Wong, K.F., Davies, D.D. (1973) Regulation of phosphoenolpyruvate carboxylase of Zea mays by metabolites. Biochem. J. 131, 451–458

    Google Scholar 

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Author notes

  1. J. W. Anderson

    Present address: Botany Department, La Trobe University, Bundoora, 3083, Victoria, Australia

Authors and Affiliations

  1. A.R.C. Research Group on Photosynthesis, Botany Department, University of Sheffield, S10 2TN, Sheffield, UK

    J. W. Anderson, Christine H. Foyer & D. A. Walker

Authors
  1. J. W. Anderson
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  2. Christine H. Foyer
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  3. D. A. Walker
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Anderson, J.W., Foyer, C.H. & Walker, D.A. Light-dependent reduction of dehydroascorbate and uptake of exogenous ascorbate by spinach chloroplasts. Planta 158, 442–450 (1983). https://doi.org/10.1007/BF00397738

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  • DOI: https://doi.org/10.1007/BF00397738

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