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Etioplasts lack thylakoid membranes and photosystem complexes. Light triggers differentiation of etioplasts into mature chloroplasts, and photosystem complexes assemble in parallel with thylakoid membrane development. Plastids isolated at various time points of de-etiolation are ideal to study the kinetic biogenesis of photosystem complexes during chloroplast development. Here, we investigated the chronology of photosystem II (PSII) biogenesis by monitoring assembly status of chlorophyll-binding protein complexes and development of water splitting via O2 production in plastids (etiochloroplasts) isolated during de-etiolation of barley ( Hordeum vulgare L.). Assembly of PSII monomers, dimers and complexes binding outer light-harvesting antenna [PSII-light-harvesting complex II (LHCII) supercomplexes] was identified after 1, 2 and 4 h of de-etiolation, respectively. Water splitting was detected in parallel with assembly of PSII monomers, and its development correlated with an increase of bound Mn in the samples. After 4 h of de-etiolation, etiochloroplasts revealed the same water-splitting efficiency as mature chloroplasts. We conclude that the capability of PSII to split water during de-etiolation precedes assembly of the PSII-LHCII supercomplexes. Taken together, data show a rapid establishment of water-splitting activity during etioplast-to-chloroplast transition and emphasize that assembly of the functional water-splitting site of PSII is not the rate-limiting step in the formation of photoactive thylakoid membranes. [ABSTRACT FROM AUTHOR]

Cyanobacteria, algae, and plants oxidize water to the O2 we breathe, and consume CO2 during the synthesis of biomass. Al- though these vital processes are functionally and structurally well separated in photosynthetic organisms, there is a long-debated role for CO2/HCO3- in water oxidation. Using membrane-inlet mass spectrometry we demonstrate that HCO3- acts as a mobile proton acceptor that helps to transport the protons produced in- side of photosystem II by water oxidation out into the chloroplast's lumen, resulting in a light-driven production of O2 and CO2. Depletion of HCO3- from the media leads, in the absence of added buffers, to a reversible down-regulation of O2 production by about 20%. These findings add a previously unidentified component to the regulatory network of oxygenic photosynthesis and conclude the more than 50-y-long quest for the function of CO2/ HCO3- in photosynthetic water oxidation. [ABSTRACT FROM AUTHOR]

Abstract: In this study we employ isotope ratio membrane-inlet mass spectrometry to probe the turnover efficiency of photosystem II (PSII) membrane fragments isolated from spinach at flash frequencies between 1Hz and 50Hz in the presence of the commonly used exogenous electron acceptors potassium ferricyanide(III) (FeCy), 2,5-dichloro-p-benzoquinone (DCBQ), and 2-phenyl-p-benzoquinone (PPBQ). The data obtained clearly indicate that among the tested acceptors PPBQ is the best at high flash frequencies. If present at high enough concentration, the PSII turnover efficiency is unaffected by flash frequency of up to 30Hz, and at 40Hz and 50Hz only a slight decrease by about 5–7% is observed. In contrast, drastic reductions of the O2 yields by about 40% and 65% were found at 50Hz for DCBQ and FeCy, respectively. Comparison with literature data reveals that PPBQ accepts electrons from QA − in PSII membrane fragments with similar efficiency as plastoquinone in intact cells. Our data also confirm that at high flashing rates O2 evolution is limited by the reactions on the electron-acceptor side of PSII. The relevance of these data to the evolutionary development of the water-splitting complex in PSII and with regard to the potential of artificial water-splitting catalysts is discussed. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: from Natural to Artificial. [Copyright &y& Elsevier]

Oxygenic photosynthesis is the basis for aerobic life on earth. The catalytic Mn4OxCaYZ center of photosystem II (PSII), after fourfold oxidation, extracts four electrons from two water molecules to yield dioxygen. This reaction cascade has appeared as a single four-electron transfer that occurs in typically 1 ms. Inevitable redox intermediates have so far escaped detection, probably because of very short lifetime. Previous attempts to stabilize intermediates by high O2-back pressure have revealed controversial results. Here we monitored by membrane-inlet mass spectrometry (MIMS) the production of 18O2 from 18O-labeled water against a high background of 6O2 in a suspension of PSII-core complexes. We found neither an inhibition nor an altered pattern of O2 production by up to 50-fold increased concentration of dissolved O2. Lack of inhibition is in line with results from previous X-ray absorption and visible-fluorescence experiments, but contradictory to the interpretation of previous UV-absorption data. Because we used essentially identical experimental conditions in MIMS as had been used in the UV work, the contradiction was serious, and we found it was not to be resolved by assuming a significant slowdown of the O2 release kinetics or a subsequent slow conformational relaxation. This calls for reevaluation of the less direct UV experiments. The direct detection of O2 release by MIMS shows unequivocally that O2 release in PSII is highly exothermic. Under the likely assumption that one H+ is released in the S4 → S0 transition, the driving force at pH 6.5 and atmospheric O2 pressure is at least 220 meV, otherwise 160 meV. [ABSTRACT FROM AUTHOR]