SITE-DIRECTED PHOTOSYSTEM-II MUTANTS WITH PERTURBED OXYGEN-EVOLVING PROPERTIES .2. INCREASED BINDING OR PHOTOOXIDATION OF MANGANESE IN THE ABSENCE OF THE EXTRINSIC 33-KDA POLYPEPTIDE IN-VIVO

Several site-directed photosystem II mutants with substitutions at Asp-170 or in the carboxyterminal region of the D1 polypeptide were characterized in vivo in the absence of the extrinsic 33-kDa polypeptide. Site-directed mutations were constructed in the cyanobacterium Synechocystis sp. PCC 6803. The 33-kDa polypeptide was removed by insertional inactivation of the Synechocystis psbO gene. Mutants were characterized by measuring changes in the yield of variable chlorophyll a fluorescence following a saturating flash or brief illumination in the presence of an electron-transfer inhibitor or following each of a series of saturating flashes in the absence of inhibitor [Chu, H.-A., Nguyen, A. P., and Debus, R. J. (1994) Biochemistry (preceding paper in this issue)]. In the presence of the extrinsic 33-kDa polypeptide, many site-directed mutants contained a significant fraction of photosystem II reaction centers that lacked photooxidizable Mn ions. This fraction decreased dramatically in the absence of the extrinsic 33-kDa polypeptide, even in mutants having a significantly perturbed high-affinity Mn binding site (e.g., in the mutants D170A and D170T). These results show that, in vivo, the extrinsic 33-kDa polypeptide directly or indirectly governs the occupancy of the high-affinity Mn binding site by Mn ions or the ability of bound Mn ions to reduce Y-Z(+)