FUNCTIONAL RECONSTITUTION OF PHOTOSYSTEM-II WITH RECOMBINANT MANGANESE-STABILIZING PROTEINS CONTAINING MUTATIONS THAT REMOVE THE DISULFIDE BRIDGE

The 33-kDa extrinsic subunit of PSII stabilizes the O-2-evolving tetra nuclear Mn cluster and accelerates O-2 evolution. We have used site-di rected mutagenesis to replace one or both Cys residues in spinach MSP with Ala. Previous experiments using native and reduced MSP led to the conclusion that a disulfide bridge between these two cysteines is ess ential both for its binding and its functional properties. We report h ere that the disulfide bridge, though essential for MSP stability, is otherwise dispensible. The mutation C51A by itself had a delayed effec t on MSP function: [C51A]MSP restored normal rates of O-2 evolution to PSII but was defective in stabilizing this activity during extended i llumination. In contrast, the Cys-free double mutant, [C28A,C51A]MSP, was functionally identical to the wild-type protein. Based on results presented here, we propose a light-dependent interaction between MSP a nd PSII that occurs only during the redox cycling of the Mn cluster an d which is destabilized by the single mutation, C51A.