Kinetics of 0₂ Evolution from H₂0₂ Catalyzed by the Oxygen-Evolving Complex: Investigation of the S₁-Dependent Reaction

The evolution of 0₂ from H₂0₂ catalyzed by the oxygen-evolving complex (OEC) in darkness was examined with photosystem II reaction center complex preparations from spinach. Flash illumination of dark-adapted reaction centers was used to make S₀-enriched or S_renriched complexes. The membranes catalyzed 0₂ evolution from H₂0₂ when preset to either the S₀or S₁state. However, only the S₀-state reaction was inhibited by carbonyl cyanide m-chlorophenylhydrazone and dependent on chloride. These results indicate that (1) the S₀-dependent and S_1-dependent catalytic cycles can be separated by flash illumination, (2) the S₀-dependent reaction involves the formation of the S₂ state, and (3) the S₁-dependent reaction does not involve the formation of the S₂ or S₃ states. A kinetic study of the S₁-dependent reaction revealed a rapid equilibrium ordered mechanism in which (1) the binding of Ca(II) must precede the binding of H₂0₂ to the OEC and (2) the reaction of Ca(II) with the free enzyme is at thermodynamic equilibrium such that Ca(II) does not necessarily dissociate after each catalytic cycle.