Calcium controls the assembly of the photosynthetic water-oxidizing complex: a cadmium(II) inorganic mutant of the Mn 4 Ca core

Abstract

Perturbation of the catalytic inorganic core (Mn ₄ Ca ₁ O _x Cl _y ) of the photosystem II-water-oxidizing complex (PSII-WOC) isolated from spinach is examined by substitution of Ca ²⁺ with cadmium(II) during core assembly. Cd ²⁺ inhibits the yield of reconstitution of O ₂ -evolution activity, called photoactivation, starting from the free inorganic cofactors and the cofactor-depleted apo-WOC-PSII complex. Ca ²⁺ affinity increases following photooxidation of the first Mn ²⁺ to Mn ³⁺ bound to the ‘high-affinity’ site. Ca ²⁺ binding occurs in the dark and is the slowest overall step of photoactivation (IM ₁ →IM ₁ ^* step). Cd ²⁺ competitively blocks the binding of Ca ²⁺ to its functional site with 10- to 30-fold higher affinity, but does not influence the binding of Mn ²⁺ to its high-affinity site. By contrast, even 10-fold higher concentrations of Cd ²⁺ have no effect on O ₂ -evolution activity in intact PSII-WOC. Paradoxically, Cd ²⁺ both inhibits photoactivation yield, while accelerating the rate of photoassembly of active centres 10-fold relative to Ca ²⁺ . Cd ²⁺ increases the kinetic stability of the photooxidized Mn ³⁺ assembly intermediate(s) by twofold (mean lifetime for dark decay). The rate data provide evidence that Cd ²⁺ binding following photooxidation of the first Mn ³⁺ , IM ₁ →IM ₁ ^* _, causes three outcomes: (i) a longer intermediate lifetime that slows IM ₁ decay to IM ₀ by charge recombination, (ii) 10-fold higher probability of attaining the degrees of freedom (either or both cofactor and protein d.f.) needed to bind and photooxidize the remaining 3 Mn ²⁺ that form the functional cluster, and (iii) increased lability of Cd ²⁺ following Mn ₄ cluster assembly results in (re)exchange of Cd ²⁺ by Ca ²⁺ which restores active O ₂ -evolving centres. Prior EPR spectroscopic data provide evidence for an oxo-bridged assembly intermediate, Mn ³⁺ (μ-O ²⁻ )Ca ²⁺ , for IM ₁ ^* . We postulate an analogous inhibited intermediate with Cd ²⁺ replacing Ca ²⁺ .