Photosynthesis Research

, Volume 110, Issue 2, pp 89–98 | Cite as

Carbonic anhydrase activity in Arabidopsis thaliana thylakoid membrane and fragments enriched with PSI or PSII

  • Lyudmila K. Ignatova
  • Natalia N. Rudenko
  • Vilen A. Mudrik
  • Tat’yana P. Fedorchuk
  • Boris N. Ivanov
Regular Paper

Abstract

The procedure of isolating the thylakoids and the thylakoid membrane fragments enriched with either photosystem I or photosystem II (PSI- and PSII-membranes) from Arabidopsis thaliana leaves was developed. It differed from the one used with pea and spinach in durations of detergent treatment and centrifugation, and in concentrations of detergent and Mg2+ in the media. Both the thylakoid and the fragments preserved carbonic anhydrase (CA) activities. Using nondenaturing electrophoresis followed by detection of CA activity in the gel stained with bromo thymol blue, one low molecular mass carrier of CA activity was found in the PSI-membranes, and two carriers, a low molecular mass one and a high molecular mass one, were found in the PSII-membranes. The proteins in the PSII-membranes differed in their sensitivity to acetazolamide (AA), a specific CA inhibitor. AA at 5 × 10−7 M inhibited the CA activity of the high molecular mass protein but stimulated the activity of the low molecular mass carrier in the PSII-membranes. At the same concentration, AA moderately inhibited, by 30%, the CA activity of PSI-membranes. CA activity of the PSII-membranes was almost completely suppressed by the lipophilic CA inhibitor, ethoxyzolamide at 10−9 M, whereas CA activity of the PSI-membranes was inhibited by this inhibitor even at 5 × 10−7 M just the same as for AA. The observed distribution of CA activity in the thylakoid membranes from A. thaliana was close to the one found in the membranes of pea, evidencing the general pattern of CA activity in the thylakoid membranes of C3-plants.

Keywords

Carbonic anhydrase Arabidopsis thaliana Thylakoids Photosystem I Photosystem II Acetazolamide Ethoxyzolamide 

Abbreviations

AA

Acetazolamide

CA

Carbonic anhydrase

Chl

Chlorophyll

DCBQ

2,6-Dichloro-p-benzoquinone

DM

Dodecyl-β-d-maltoside

EZ

Ethoxyzolamide

PAAG

Polyacrylamide gel

PMSF

Phenylmethylsulfonylfluoride

PSI

Photosystem I

PSII

Photosystem II

Rubisco

Ribulosebisphosphatecarboxylase/oxygenase

TMPD

N,N,N′,N′-tetramethyl-p-phenilene diamine

Notes

Acknowledgments

The authors express their gratitude to Dr. S. Khorobrykh for his help in the study, and to I. Naydov for the data of confocal microscope image. This study was supported by the grant from Civilian Research & Development Foundation (CRDF) (project RUB1-2911-PU-07) and Russian Foundation for Basic Research (RFBR) (project 08-04-00480_a).

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Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  • Lyudmila K. Ignatova
    • 1
  • Natalia N. Rudenko
    • 1
  • Vilen A. Mudrik
    • 1
  • Tat’yana P. Fedorchuk
    • 1
  • Boris N. Ivanov
    • 1
  1. 1.Institute of Basic Biological Problems of Russian Academy of SciencesPushchino, MoscowRussia

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