Abstract
Photosynthetic O(2) evolution is catalyzed by the Mn(4)CaO(5) cluster of the water oxidation complex of the photosystem II (PSII) complex. The photooxidative self-assembly of the Mn(4)CaO(5) cluster, termed photoactivation, utilizes the same highly oxidizing species that drive the water oxidation in order to drive the incorporation of Mn(2+) into the high-valence Mn(4)CaO(5) cluster. This multistep process proceeds with low quantum efficiency, involves a molecular rearrangement between light-activated steps, and is prone to photoinactivation and misassembly. A sensitive polarographic technique was used to track the assembly process under flash illumination as a function of the constituent Mn(2+) and Ca(2+) ions in genetically engineered membranes of the cyanobacterium Synechocystis sp. PCC6803 to elucidate the action of Ca(2+) and peripheral proteins. We show that the protein scaffolding organizing this process is allosterically modulated by the assembly protein Psb27, which together with Ca(2+) stabilizes the intermediates of photoactivation, a feature especially evident at long intervals between photoactivating flashes. The results indicate three critical metal-binding sites: two Mn and one Ca, with occupation of the Ca site by Ca(2+) critical for the suppression of photoinactivation. The long-observed competition between Mn(2+) and Ca(2+) occurs at the second Mn site, and its occupation by competing Ca(2+) slows the rearrangement. The relatively low overall quantum efficiency of photoactivation is explained by the requirement of correct occupancy of these metal-binding sites coupled to a slow restructuring of the protein ligation environment, which are jointly necessary for the photooxidative trapping of the first stable assembly intermediate.