RELATIONSHIP BETWEEN ACTIVITY, D1 LOSS, AND MN BINDING IN PHOTOINHIBITION OF PHOTOSYSTEM-II

Photoinhibition of photosystem II (PSII) activity and loss of the D1 r eaction center protein were studied in PSII-enriched membrane fragment s in which the water-splitting complex was inhibited by depletion of e ither calcium or chloride or by removing manganese. The Ca2+-depleted PSII was found to be the least susceptible to inhibition by light as r eported previously (Krieger, A., and Rutherford, A. W. (1997) Biochim. Biophys. Acta 1319, 91-98). This different susceptibility to light wa s not reflected in the extent of D1 protein loss. In Mn-depleted PSII the loss of activity and the loss of the D1 protein were correlated, w hile in Cl-- and Ca2+-depleted PSII, there was very little loss of the D1 protein. The production of free radicals and singlet oxygen was me asured by EPR spin-trapping techniques in the different samples. O-1(2 ) and carbon-centered radicals could be detected after photoinhibition of active PSII, while hydroxyl radical formation dominated in all of the other samples. In addition, photoinhibition of PSII was investigat ed in which the functional Mn cluster was reconstituted (i.e., photoac tivated). As expected this led to a protection against photoinhibition . When the photoactivation procedure was done in the absence of Ca2+ n o activity was obtained although a nonfunctional Mn cluster was formed . Despite the lack of activity the binding of Mn partially protected a gainst the loss of D1. These data demonstrate that, during photoinhibi tion, the extent of D1 loss is neither affected by the water-splitting activity of the sample nor correlated to the kinetics of PSII activit y loss. D1 loss seems to be independent of the chemical nature of the reactive oxygen species formed during photoinhibition and seems to occ ur only in the absence of Mn. It is proposed that Mn binding protects against D1 loss by maintaining a protein structure which is not access ible to cleavage.