Skip to main content
SpringerLink
Log in

Loose association of ribulose 1,5-bisphosphate carboxylase/oxygenase with chloroplast thylakoid membranes

  • Published:
Photosynthesis Research Aims and scope Submit manuscript

Abstract

The intra-chloroplastic distribution of ribulose 1,5-bisphosphate carboxylase/oxygenase (RuBisCO) between thylakoid membranes and stroma was studied by determining the enzyme activities in the two fractions, obtained by the rapid centrifugation of hypotonically disrupted chloroplast preparations of spinach and pea leaf tissues. The membrane-associated form of RuBisCO was found to increase in proportion to the concentration of MgCl2 in the disrupting medium; with 20 mM MgCl2 approximately 20% of the total RuBisCO of spinach chloroplasts and 10% of that of pea chloroplasts became associated with thylakoid membranes. Once released from membranes in the absence of MgCl2, addition of MgCl2 did not cause reassociation of the enzyme. The inclusion of KCl in the hypotonic disruption buffer also caused the association of RuBisCO with membranes; however, up to 30 mM KCl, only minimal enzyme activities could be detected in the membranes, whereas above 40 mM KCl there was a sharp increase in the membrane-associated form of the enzyme.

Higher concentrations of chloroplasts during the hypotonic disruption, as well as addition of purified preparations of RuBisCO to the hypotonic buffer, resulted in an increase of membrane-associated activity. Therefore, the association of the enzyme with thylakoid membranes appears to be dependent on the concentration of RuBisCO. P-glycerate kinase and aldolase also associated to the thylakoid membranes but NADP-linked glyceraldehyde-3-P dehydrogenase did not. The optimal conditions for enzyme association with the thylakoid membranes were examined; maximal association occurred at pH 8.0. The association was temperature-insensitive in the range of 4° to 25° C. RuBisCO associated with the thylakoid membranes could be gradually liberated to the soluble form upon shaking in a Vortex mixer at maximal speed, indicating that the association is loose.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

Abbreviations

DTT:

dithiothreitol

RuBP:

ribulose 1,5-bisphosphate

RuBisCO:

ribulose 1,5-bisphosphate carboxylase/oxygenase

MES:

2-(N-morpholino) ethane sulfonic acid

References

  1. Akazawa T (1977) Structure and function of ribulose bisphosphate carboxylase in Proceedings IVth Int. Cong. Photosynthesis. (Hall, D.O., Coombs, J. and Goodwin, T.W., eds.). The biochemical Society 447–456

  2. Alscher-Herman R (1982) Chloroplast alkaline fructose 1,6-bisphosphatase exists in a membrane-bound form. Plant Physiol 70: 728–734

    Google Scholar 

  3. Anderson LE and Ben-Bassat D (1981) Membrane-bound carbon metabolism enzymes in pea leaf chloroplasts in Proceedings Vth Int. Congr. Photosynthesis. IV (G. Akoyunoglou ed.). Balaban International Science Services 373–378

  4. Anderson LE, Ng TL, and Park KY (1974) Inactivation of pea leaf chloroplastic and cytoplasmic glucose 6-phosphate dehydrogenase by light and dithiothreitol. Plant Physiol 53: 835–839

    Google Scholar 

  5. Badger MR and Lorimer GH (1981) Interaction of sugarphosphates with the catalytic site of ribulose 1,5-bisphosphate carboxylase. Biochemistry 20: 2219–2225

    Google Scholar 

  6. Bahr JT and Jensen RG (1978) Activation of ribulose bisphosphate carboxylase in intact chloroplasts by CO2 and light. Arch Biochem Biophys 185: 39–48

    Google Scholar 

  7. Barber J (1980) Membrane surface charges and potentials in relation to photosynthesis. Biochim Biophys Acta 594: 253–308

    Google Scholar 

  8. Cockburn W, Walker DA and Baldry CW (1968) The isolation of spinach chloroplasts in pyrophosphate media. Plant Physiol 43: 1415–1418

    Google Scholar 

  9. Garrett MK (1978) Control of photorespiration at RuBP carboxylase/oxygenase evel in rye grass cultivars. Nature 274: 913–915

    Google Scholar 

  10. Fischer KH and ELatzko (1979) Chloroplast ribulose-5-phosphate kinase: light-mediated activation, and detection of both soluble and membrane associated activity. Biochem Biophys Res Commun 89: 300–306

    Google Scholar 

  11. Heber Ü, and Santarius KA (1970) Direct and indirect transport of ATP and ADP across the chloroplast envelope 2. Naturforsch 256: 718–728

    Google Scholar 

  12. Heldt HW, Chon C Ja and Lorimer GH (1978) Phosphate requirement for the light activation of ribulose 1,5-bisphosphate carboxylase in intact spinach chloroplasts. FEBS Lett 92: 234–240

    Google Scholar 

  13. Heldt HW, Werdan K, Milovancev M and Geller G (1973) Alkalization of the chloroplast stroma caused by light dependent proton flux into the thylakoid space. Biochem Biophys Acta 314: 224–241

    Google Scholar 

  14. Henriques A and Park RB (1976) Identification of chloroplast membrane peptides with subunits of coupling factor and ribulose 1,5-diphosphate carboxylase. Arch Biochem Biophys 176: 472–478

    Google Scholar 

  15. Howell SH and Moudrianakis EN (1967) Function of the ‘quantasome’ in photosynthesis: Structure and properties of membrane-bound particle active in the dark reactions of photophosphorylation. Proc Natl Acad Sci 58: 1261–1268

    Google Scholar 

  16. Kannangara CG, vanWyk D and Menke W (1970) Immunological evidence for the presence of latent Ca2+ dependent ATPase and carboxydismutase on the thylakoid surface. Z. Naturforsch 25B: 613–618

    Google Scholar 

  17. Krause GH (1977) Light-induced movement of magnesium ions in intact chloroplasts. Spectroscopic determination with eriochrome blue SE. Biochim Biophys Acta 460: 500–510

    Google Scholar 

  18. Leegood RC and Walker DA (1980) Autocatalysis and light activation of enzymes in relation to photosynthetic induction in wheat chloroplasts. Arch Biochem Biophys 200: 575–582

    Google Scholar 

  19. Lendzian KJ (1978) Activation of ribulose 1,5-bisphosphate carboxylase by chloroplast metabolites in a reconstituted spinach chloroplast system. Planta 143: 291–296

    Google Scholar 

  20. McNeil PH and Walker DA (1981) The effect of magnesium and other ions on the distribution of ribulose 1,5-bisphosphate carboxylase in chloroplast extracts. Arch Biochem Biophys 208: 184–188

    Google Scholar 

  21. Miller KR and Staehlin LA (1976) Analysis of the thylakoid outer surface coupling factor is limited to unstacked membrane regions. J Cell Biol 68: 30–47

    Google Scholar 

  22. Pacold I and Anderson LE (1975) Chloroplast and cytoplasmic enzymes. Plant Physiol 55: 168–171

    Google Scholar 

  23. Perchorowicz JT, Raynes DA and Jensen RG (1981) Light limitation of photosynthesis and activation of ribulose bisphosphate carboxylase in wheat seedlings. Proc Natl Acad Sci 78: 2985–2989

    Google Scholar 

  24. Portis AR and Heldt HW (1976) Light dependent changes of the Mg2+ concentration in the stroma in relation to the Mg2+ dependency of CO2 fixation in intact chloroplasts. Biochim Biophys Acta 449: 434–446

    Google Scholar 

  25. Raghavendra AS, Carrillo N and Vallejos RH (1981) The pattern and characteristics of membrane bound Ribulose bisphosphate carboxylase in spinach chloroplasts in proceedings Vth Int. Congr. Photosynthesis. IV (G. Akoyunoglou ed). Balaban International Science Services 31–38

  26. Robinson SP, McNeil PH and Walker DA (1979) Ribulose bisphosphate carboxylase-lack of dark inactivation of the enzyme in experiments with protoplasts. FEBS Lett 97: 296–300

    Google Scholar 

  27. Scheibe R and Beck E (1979) On the mechanism of activation by light of the NADP dependent malate dehydrogenase in spinach chloroplasts. Plant Physiol 64: 744–748

    Google Scholar 

  28. Shahak Y (1982) The role of Mg2+ in the light activation process of the H+-ATPase in intact chloroplasts. FEBS Lett 145: 223–229

    Google Scholar 

  29. Strotmann H, Hesse H and Edelman K (1973) Quantitative determination of coupling factor CF1 of chloroplasts. Biochim Biophys Acta 314: 202–210

    Google Scholar 

  30. Takabe T, Nishimura M and akazawa T (1976) Presence of two subunit types in ribulose 1,5-bisphosphate carboxylase from blue green algae. Biochem Biophys Res Commun 68: 537–544

    Google Scholar 

  31. Takabe T, Nishimura M and Akazawa T (1979) Isolation of intact chloroplasts from spinach leaf by centrifugation in gradients of the modified silica ‘Percoll’. Agric Biol Chem 43: 2137–2142

    Google Scholar 

  32. Wu R and Racker E (1959) Regulatory mechanisms in carbohydrate metabolism. III Limiting factors in glycolysis of ascites tumor cells. J Biol Chem 234: 1029–1035

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Research Institute for Biochemical Regulation, School of Agriculture, Nagoya University, 464, Chikusa, Nagoya, Japan

    H. Mori, Tetsuko Takabe & T. Akazawa

Authors
  1. H. Mori
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Tetsuko Takabe
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. T. Akazawa
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Mori, H., Takabe, T. & Akazawa, T. Loose association of ribulose 1,5-bisphosphate carboxylase/oxygenase with chloroplast thylakoid membranes. Photosynth Res 5, 17–28 (1984). https://doi.org/10.1007/BF00018372

Download citation

  • Received:

  • Revised:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00018372

Keywords

Search

Navigation