Skip to main content

Measurement of the ΔpH and Electric Field Developed Across Arabidopsis Thylakoids in the Light

  • Protocol
  • First Online:
Chloroplast Research in Arabidopsis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 775))

Abstract

Measurement of the different components of the proton motive force (pmf) gives information about the coupling of proton movement within thylakoids to chemiosmotic processes such as photophosphorylation and protein transport, as well as that relating to the overall quality of a thylakoid preparation. The techniques to assess the pmf have been known for many years, as they have been applied to the most popular model plants for photosynthetic research. The emergence of Arabidopsis thaliana as the prominent model plant in developmental and genetics research prompted us to apply these techniques to thylakoids isolated from Arabidopsis chloroplasts. We describe here two spectroscopic techniques to measure the transmembrane pH gradient and electric field developed in the light in Arabidopsis thylakoids.

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

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 159.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Similar content being viewed by others

References

  1. Mitchell, P. (1966) Chemiosmotic coupling in oxidative and photosynthetic phosphorylation. Biological Review 41, 445–502.

    Article  CAS  Google Scholar 

  2. Alder, N. N., and Theg, S. M. (2003) Energetics of protein transport across biological membranes: A study of the thylakoid DeltapH-dependent/cpTat pathway. Cell 112, 231–242.

    Article  PubMed  CAS  Google Scholar 

  3. Braun, N. A., Davis, A. W., and Theg, S. M. (2007) The chloroplast tat pathway utilizes the transmembrane electric potential as an energy source. Biophys. J. 93, 1993–1998.

    Article  PubMed  CAS  Google Scholar 

  4. Teter, S. A., and Theg, S. M. (1998) Energy-transducing thylakoid membranes remain highly impermeable to ions during protein translocation. Proc. Natl. Acad. Sci. USA 95, 1590–1594.

    Article  PubMed  CAS  Google Scholar 

  5. Theg, S. (2010) Measurement of the energetics of protein transport across the chloroplast thylakoid membrane. Methods Mol. Biol. 619, 323–337.

    Article  PubMed  CAS  Google Scholar 

  6. Schuldiner, S., Rottenberg, H., and Avron, M. (1972) Determination of delta pH in chloroplasts: 2. Fluorescent amines as a probe for the determination of delta pH in chloroplasts. Eur. J. Biochem. 25, 64–70.

    CAS  Google Scholar 

  7. Mills, J. (1986) Photophosphorylation. In, Photosynthesis, Energy Transduction: A Practical Approach (Hipkins, M., and Baker, N., eds.) IRL Press, Washington, D.C., USA, pp. 143–187.

    Google Scholar 

  8. Junge, W., and Witt, H. T. (1968) On the ion transport system of photosynthesis - investigations on a molecular level. Z. Naturforsch. 23, 244–254.

    CAS  Google Scholar 

  9. Sacksteder, C. A., Jacoby, M. E., and Kramer, D. M. (2001) A portable, non-focusing optics spectrophotometer (NoFOSpec) for measurements of steady-state absorbance changes in intact plants. Photosynth Res. 70, 231–240.

    Article  PubMed  CAS  Google Scholar 

  10. Junge, W. (1976) Flash kinetic spectroscopy in the study of plant pigments. In, Biochemistry of Plant Pigments, Vol. 11 (Goodwin, T., ed.) Academic Press, New York, USA, pp. 233–333.

    Google Scholar 

  11. Berry, S., and Rumberg, B. (1999) Proton to electron stoichiometry in electron transport of spinach thylakoids. Biochim. Biophys. Acta 1410, 248–61.

    Article  PubMed  CAS  Google Scholar 

  12. Walz, D., Goldstein, L., and Avron, M. (1974) Determination and analysis of the buffer capacity of isolated chloroplasts in the light and in the dark. Eur. J. Biochem. 47, 403–407.

    Article  PubMed  CAS  Google Scholar 

  13. Junge, W., Auslander, W., McGeer, A. J., and Runge, T. (1979) The buffering capacity of the internal phase of thylakoids and the magnitude of the pH changes inside under flashing light. Biochim. Biophys. Acta 546, 121–141.

    Article  PubMed  CAS  Google Scholar 

  14. Junge, W. (1982) Electrogenic reactions and proton pumping in green plant photosynthesis. In, Current Topics in Membranes and Transport, Vol. 16 (Slayman, C. L., ed.) Academic Press, New York, USA, pp. 431–464.

    Google Scholar 

Download references

Acknowledgments

This work was supported by US Department of Energy Grant DE-FG02-03ER15405.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Steven M. Theg .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2011 Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Theg, S.M., Tom, C. (2011). Measurement of the ΔpH and Electric Field Developed Across Arabidopsis Thylakoids in the Light. In: Jarvis, R. (eds) Chloroplast Research in Arabidopsis. Methods in Molecular Biology, vol 775. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-237-3_18

Download citation

  • DOI: https://doi.org/10.1007/978-1-61779-237-3_18

  • Published:

  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-61779-236-6

  • Online ISBN: 978-1-61779-237-3

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics