Stoichiometry, inhibitor sensitivity, and organization of manganese associated with photosynthetic oxygen evolution

Chloroplast thylakoid membranes isolated in the presence of EDTA retain high rates of O(2) evolution (>/=340 mumol.h(-1).mg chlorophyll(-1)) but contain no Mn(2+) that is detectable by electron paramagnetic resonance (EPR) at room temperature. The total Mn(2+) content of these preparations is 4.6 per 400 chlorophylls; 0.6 Mn(2+) can be released by addition of Ca(2+), a treatment that does not affect O(2) evolution. The remaining Mn(2+) (4 per 400 chlorophylls) appears to be functionally associated with O(2) evolution activity. Inhibition by Tris, NH(2)OH, or heat will release a small fraction of Mn(2+) from these membranes ( approximately 25% with Tris, for example). Addition of Ca(2+) further enhances Mn(2+) release so that for Tris and for NH(2)OH, 2 and 3, respectively, Mn(2+) per 400 chlorophylls are extracted from the O(2)-evolving complex. Based on the microwave power-saturation properties of the EPR signal IIf, which arises from an intermediate electron carrier in the water splitting process, it appears that one of the four Mn(2+) associated with photosystem II is uniquely sensitive to Tris. A new model is proposed for the organization and inhibitor sensitivity of manganese in the O(2)-evolving complex.