It was previously shown in the photosystem II membrane preparation DT- 20 that photoxidation of the oxygen-evolving manganese cluster was blocked by 0.1 mM formate, unless 0.2 mM bicarbonate was present as well [Wincenejusz, H., Allakhverdiev. S. I., Klimov, V. V., and Van Gorkom, H. J. (1996) Biochim. Biophys. Acta 1273, 1-3]. Here it is shown by measurements of EPR signal II that oxidation of the secondary electron donor, Y(Z), is not inhibited. However, the reduction of Y+/(Z) is greatly slowed and occurs largely by back reaction with reduced acceptors. Bicarbonate is shown to prevent the loss of fast electron donation to Y+(Z). The release of about one or two free Mn2+ per photosystem II during formate treatment, and the fact that these effects are mimicked by Mn-depletion, suggests that formate may act by replacing a bicarbonate which is essential for Mn binding. Irreversible light-induced rebinding in an EPR-silent form of Mn2+ that was added to Mn-depleted DT-20 was indeed found to depend on the presence of bicarbonate, as did the reconstitution in such material of both the fast electron donation to Y+/(Z) and the UV absorbance changes characteristic of a functional oxygen-evolving complex. It is concluded that bicarbonate may be an essential ligand of the functional Mn cluster.
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