Evidence for an association between a 33 kDa extrinsic membrane protein, manganese and photosynthetic oxygen evolution. I. Correlation with the S2 multiline EPR signal

D. Hunziker, D. R. Abramowicz, R. Damoder, G Charles Dismukes

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The removal of peripheral membrane proteins of a molecular mass of 17 and 23 kDa by washing of spinach Photosystem-II (PS II) membranes in 1 M salt between pH 4.5 and 6.5 produces a minimal loss of the S1 → S2 reaction, as seen by the multiline EPR signal for the S2 state of the water-oxidizing complex, while reversibly inhibiting O2 evolution. The multiline EPR signal simplifies from a '19-line' spectrum to a '16-line' spectrum, suggestive of partial uncoupling of a cluster of 3 or 4 to yield photo-oxidation of a binuclear Mn site. Alkaline salt washing progressively releases a 33 kDa peripheral protein between pH 6.5 and 9.5, in direct parallel with the loss of O2 evolution and the S2 multiline EPR signal. The 33 kDa protein can be partially removed (20%) at pH 8.0 prior to managanese release. Salt treatment releases four Mn ions between pH 8.0 and 9.5 with the first 2 or 3 Mn ions released cooperatively. A common binding site is thus suggested in agreement with earlier EPR spectroscopic data establishing a tetranuclear Mn site. At least two of these Mn ions bind directly at a site in the PS II complex for which photooxidation by the reaction center is controlled by the 33 kDa protein. The washing of PS II membranes with 1 M CaCl2 to affect the release of the 33 kDa protein, while preserving Mn binding to the membrane (Ono, T.-A. and Inoue, Y. (1983) FEBS Lett. 164, 255-260), is found to leave some 33 kDa protein undissociated in proportion to the extent of O2 evolution and S2 multiline yield. These depleted membranes do not oxidize water or produce the normal S2 state without the binding of the 33 kDa protein. A method for the accurate determination of relative concentrations of the peripheral membrane proteins using gel electrophoresis is presented.

Original languageEnglish
Pages (from-to)6-14
Number of pages9
JournalBiochimica et Biophysica Acta - Bioenergetics
Issue number1
Publication statusPublished - Jan 16 1987



  • (EPR)
  • Manganese
  • Oxygen evolution
  • Photosynthesis
  • Photosystem II
  • Water oxidation

ASJC Scopus subject areas

  • Biophysics

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