Photosystem II oxygen-evolving complex photoassembly displays an inverse H/D solvent isotope effect under chloride-limiting conditions

Abstract

Photosystem II (PSII) performs the solar-driven oxidation of water used to fuel oxygenic photosynthesis. The active site of water oxidation is the oxygen-evolving complex (OEC), a Mn4CaO5 cluster. PSII requires degradation of key subunits and reassembly of the OEC as frequently as every 20 to 40 min. The metals for the OEC are assembled within the PSII protein environment via a series of binding events and photochemically induced oxidation events, but the full mechanism is unknown. A role of proton release in this mechanism is suggested here by the observation that the yield of in vitro OEC photoassembly is higher in deuterated water, D2O, compared with H2O when chloride is limiting. In kinetic studies, OEC photoassembly shows a significant lag phase in H2O at limiting chloride concentrations with an apparent H/D solvent isotope effect of 0.14 ± 0.05. The growth phase of OEC photoassembly shows an H/D solvent isotope effect of 1.5 ± 0.2. We analyzed the protonation states of the OEC protein environment using classical Multiconformer Continuum Electrostatics. Combining experiments and simulations leads to a model in which protons are lost from amino acid that will serve as OEC ligands as metals are bound. Chloride and D2O increase the proton affinities of key amino acid residues. These residues tune the binding affinity of Mn2+/3+ and facilitate the deprotonation of water to form a proposed μ-hydroxo bridged Mn2+Mn3+ intermediate.

The role of Ca2+and protein scaffolding in the formation in nature’s water oxidizing complex

“…Calcium lowers the pKa for water ligand, which is controlled by a nearby base Bthat serves as a primary proton acceptor with a pKa dependent on Ca 2+ bound to its effector site (23). The deprotonation of the intermediates and the tuning of pKas facilitated by Clbinding is critical for assembly (24). The absence of Ca 2+ during photoactivation leads to the formation of non-catalytic, multinuclear high valent Mn species ("inappropriately bound Mn") that inactivates the PSII complex (22).…”

Light-driven formation of high-valent manganese oxide by photosystem II supports evolutionary role in early bioenergetics

“…Cheniae et al investigated photoactivation and observed light-driven binding of about 18 membrane-bound Mn ions per PSII if Ca ions were excluded from the photoassembly buffer 48 , whereas we here observe binding of clearly more Mn ions (65±19 Mn ions, Table 1), irrespective of the absence or presence of Ca ions at moderate concentration (5 mM) in the photoassembly buffer ( Figure S12). The presence of 5 mM CaCl2 allows for photoactivation, although higher concentration are required for optimal photoactivation yield 48,49 . The about 20 times higher light intensities we used likely promoted the oxidation and binding of numerous Mn ions at the expense of formation of a single native Mn4CaO5 cluster, because the latter requires low light intensities presumably due to the presence of a slow 'dark rearrangement' step for assembly 19 .…”

Chloride involvement in the synthesis, functioning and repair of the photosynthetic apparatusin vivo

“…5 of Vinyard et al . (2019) showed that O 2 evolution was 20 μmol mg −1 Chl h −1 with 100 mol Cl − m −3 and 9 μmol mg −1 Chl h −1 with 20 mol Cl − m −3 . The data for PSII preparations with the psbO subunit are much more likely to represent the in vivo Cl − dependence of PSII‐OEC assembly, with a half‐saturation value for Cl − of c .…”

“…Assembly of the PSII complex, including the OEC, is essential for the growth of oxygenic photolithotrophs, and for the replacement of photodamaged PSII (Vinyard et al , 2019; Weisz et al , 2019). Focusing on the anomalous isotope effect with faster reaction with 2 H 2 O rather than 1 H 2 O as solvent, Vinyard et al .…”

“…Turning to estimates of chloroplast Cl − concentrations, many of the estimates of Cl − requirements for PSII (re‐)assembly (Vinyard et al , 2019) and function (Lindberg & Andréasson, 1996; Murray et al , 2008; Kawakami et al , 2009; Umena et al , 2011; Reiss et al , 2019) were obtained for S. oleracea , a plant for which relevant data on chloroplast Cl − concentrations are available. Although S. oleracea is tolerant of moderate salinisation, it grows well in nonsaline media.…”