ENDOR study on the position of hydrogens close to the manganese cluster in S₂ state of photosystem II

Proton matrix ENDOR of the manganese multiline in the S 2 , state of photosystem II membranes from spinach has been investigated. The spectral structure over a range of frequencies of ± 2 MHz centered on the position for a free proton was analyzed through employing a simulation method based on the dipolar interaction between electron and proton magnetic moments. The 6 pairs of lines resolved were attributed to the surrounding proton populations at distances varying within the range 2.7–6.0 Å from the putative manganese center. Two of the 6 pairs, namely those corresponding to protons at distances of 2.7 and 3.2 Å from the manganese center were eliminated on washing of the membranes with deuterated buffer. This suggests that these protons belong to the water molecules coordinating to the manganese cluster. The presence of ethylene glycol instead of glycerol and sucrose in the buffer broadened both EPR and ENDOR spectra. This suggests that ethylene glycol molecules are also accessible to the two‐manganese cluster. A possible model of water association to the manganese ions in the photosynthetic water‐oxidation system is proposed.