, Volume 56, Issue 1, pp 210–216 | Cite as

Quantification of bound bicarbonate in photosystem II

  • K. Tikhonov
  • D. ShevelaEmail author
  • V. V. Klimov
  • J. Messinger


In this study, we presented a new approach for quantification of bicarbonate (HCO3) molecules bound to PSII. Our method, which is based on a combination of membrane-inlet mass spectrometry (MIMS) and 18O-labelling, excludes the possibility of “non-accounted” HCO3 by avoiding (1) the employment of formate for removal of HCO3 from PSII, and (2) the extremely low concentrations of HCO3/CO2 during online MIMS measurements. By equilibration of PSII sample to ambient CO2 concentration of dissolved CO2/HCO3, the method ensures that all physiological binding sites are saturated before analysis. With this approach, we determined that in spinach PSII membrane fragments 1.1 ± 0.1 HCO3 are bound per PSII reaction center, while none was bound to isolated PsbO protein. Our present results confirmed that PSII binds one HCO3 molecule as ligand to the non-heme iron of PSII, while unbound HCO3 optimizes the water-splitting reactions by acting as a mobile proton shuttle.

Additional key words

hydrogen carbonate inorganic carbon mass spectrometry Mn-stabilizing protein non-heme iron oxygen-evolving complex 





membrane-inlet mass spectrometry


oxygen-evolving complex






reaction center


non-heme iron


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Copyright information

© The Institute of Experimental Botany 2018

Authors and Affiliations

  • K. Tikhonov
    • 1
  • D. Shevela
    • 2
    Email author
  • V. V. Klimov
    • 1
  • J. Messinger
    • 2
    • 3
  1. 1.Institute of Basic Biological ProblemsRussian Academy of SciencePushchinoRussia
  2. 2.Department of Chemistry, Chemical Biological CentreUmeå UniversityUmeåSweden
  3. 3.Department of Chemistry, Molecular BiomimeticsÅngström Laboratory, Uppsala UniversityUppsalaSweden

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