Bicarbonate Coordinates to Mn3+ during Photo-Assembly of the Catalytic Mn4Ca Core of Photosynthetic Water Oxidation: EPR Characterization

Abstract

Assembly of the catalytic cluster, Mn4CaOxCly, comprising the water-oxidizing complex (WOC) of photosystem II (PSII), occurs during biogenesis in the presence of the apo-WOC-PSII complex, Mn2+, Ca2+ and Cl− cofactors under weak illumination. The in vitro assembly process known as photo-activation involves several intermediates that have been resolved in previous kinetic studies. (Bi)carbonate has been shown to stimulate the rate of formation and yield of the first stable light-induced Mn3+ assembly intermediate (IM1) from Mn2+ bound to the high-affinity assembly site in apo-WOC-PSII. 13C electron spin echo envelope modulation has previously revealed that (bi)carbonate is a ligand to this Mn2+. Herein, we use parallel-mode electron paramagnetic resonance (EPR) spectroscopy to characterize the Mn3+ photoproduct, which exists as a ternary complex with carbonate at the high-affinity assembly site (in the absence of Ca2+) formulated as [CO3-Mn3+-apo-WOC-PSII]. The EPR-derived spectral parameters of IM1 (the g value, 55Mn hyperfine coupling constant (AZ) and the ligand-field splitting parameters D/E) are independent of solution pH, in marked contrast to their strong pH dependence in the absence of bicarbonate. (Bi)carbonate coordination “chemically isolates” the IM1 from external pH changes, much like that caused by Ca2+ coordination, revealing similar roles in photo-assembly. The cumulative results reveal that (bi)carbonate and Ca2+ coordination control the ligand field strength and symmetry around the initial high-affinity Mn3+, consistent with the possible formation of a μ2-oxide bridge in IM1, [Mn3+(O2−)Ca2+]. These events greatly improve the quantum yield of subsequent steps in photo-assembly.