Periodic changes in the oxidation state of manganese in photosynthetic oxygen evolution upon illumination with flashes.

The pattern of manganese released from chloroplast membranes by a rapid temperature shock after various illumination regimes indicates that changes in the oxidation state of bound manganese occur during photosynthesis. Continuous illumination decreases by 35-40% the amount of Mn(II) released in the presence of K3Fe(CN)6 compared with a dark-adapted control. Following illumination and heat treatment, the addition of the reductant H2O2 to the samples causes an increase in the level of electron paramagnetic resonance (EPR)-detectable manganese. The pH dependence of the H2O2 reduction indicates that the non-EPR-detectable manganese present in the heated sample after illumination is in the form of higher oxidation state compounds, e.g. MnO2. The light-induced Mn(II) decrease is reversible in the dark with t 1/2 approx. 40 s and can be prevented by the presence of the Photosystem II inhibitors 3-(3,4-dichlorophenyl)-1,1-dimethyl urea or fluorocarbonylcyanide phenylhydrazone during the illumination period. After a series of brief flashes of light the Mn(II) released by heat treatment oscillates over periods of four flashes. The pattern is similar to the O2 yield flash pattern and suggests that a cycling of manganese oxidation states is involved in the O2 evolution mechanism. The oscillations in the Mn(II) release are analyzed in terms of the current four-step model for O2 evolution. The analysis suggests that manganese is successively oxidized in the first two steps, but undergoes a partial reduction on the third step. This result is consistent with the concept that water undergoes a partial oxidation prior to the release of O2 from the water-splitting complex.