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Green organisms evolve oxygen (O2) via photosynthesis and consume it by respiration. Generally, net O2 consumption only becomes dominant when photosynthesis is suppressed at night. Here, we show that green thylakoid membranes of Scots pine (Pinus sylvestris L) and Norway spruce (Picea abies) needles display strong O2 consumption even in the presence of light when extremely low temperatures coincide with high solar irradiation during early spring (ES). By employing different electron transport chain inhibitors, we show that this unusual light-induced O2 consumption occurs around photosystem (PS) I and correlates with higher abundance of flavodiiron (Flv) A protein in ES thylakoids. With P700 absorption changes, we demonstrate that electron scavenging from the acceptor-side of PSI via O2 photoreduction is a major alternative pathway in ES. This photoprotection mechanism in vascular plants indicates that conifers have developed an adaptative evolution trajectory for growing in harsh environments.

In general, the word, photosynthesis, is considered synonymous with oxygenic photosynthesis, a process by which cyanobacteria, algae, aquatic, and terrestrial plants produce oxygen and carbohydrates, using light (photons), water and carbon dioxide. Further, we have anoxygenic bacterial photosynthesis where oxygen is not evolved, but a substrate, other than water, is oxidized, and rhodopsin-type systems, where ATP is produced. In principle, one could expand the concept of the term photosynthesis, provided appropriate caveats are added, to include lightdriven assimilation of molecular nitrogen, photoproduction of molecular hydrogen, and even synthesis of vitamin D in skin. We conclude with a glimpse of the rapidly developing field of artificial photosynthesis.

Green organisms evolve oxygen (O2) via photosynthesis and consume it by respiration. Generally, net O2 consumption only becomes dominant when photosynthesis is suppressed at night. Here, we show that green thylakoid membranes of Scots pine (Pinus sylvestris L) and Norway spruce (Picea abies) needles display strong O2 consumption even in the presence of light when extremely low temperatures coincide with high solar irradiation during early spring (ES). By employing different electron transport chain inhibitors, we show that this unusual light-induced O2 consumption occurs around photosystem (PS) I and correlates with higher abundance of flavodiiron (Flv) A protein in ES thylakoids. With P700 absorption changes, we demonstrate that electron scavenging from the acceptor-side of PSI via O2 photoreduction is a major alternative pathway in ES. This photoprotection mechanism in vascular plants indicates that conifers have developed an adaptative evolution trajectory for growing in harsh environments.

Photosynthetic water oxidation by Photosystem II (PSII) is a fascinating process because it sustains life on Earth and serves as a blue print for scalable synthetic catalysts required for renewable energy applications. The biophysical, computational, and structural description of this process, which started more than 50 years ago, has made tremendous progress over the past two decades, with its high-resolution crystal structures being available not only of the dark-stable state of PSII, but of all the semi-stable reaction intermediates and even some transient states. Here, we summarize the current knowledge on PSII with emphasis on the basic principles that govern the conversion of light energy to chemical energy in PSII, as well as on the illustration of the molecular structures that enable these reactions. The important remaining questions regarding the mechanism of biological water oxidation are highlighted, and one possible pathway for this fundamental reaction is described at a molecular level.

Green organisms evolve oxygen (O2) via photosynthesis and consume it by respiration. Generally, net O2 consumption only becomes dominant when photosynthesis is suppressed at night. Here, we show that green thylakoid membranes of Scots pine (Pinus sylvestris L) and Norway spruce (Picea abies) needles display strong O2 consumption even in the presence of light when extremely low temperatures coincide with high solar irradiation during early spring (ES). By employing different electron transport chain inhibitors, we show that this unusual light-induced O2 consumption occurs around photosystem (PS) I and correlates with higher abundance of flavodiiron (Flv) A protein in ES thylakoids. With P700 absorption changes, we demonstrate that electron scavenging from the acceptor-side of PSI via O2 photoreduction is a major alternative pathway in ES. This photoprotection mechanism in vascular plants indicates that conifers have developed an adaptative evolution trajectory for growing in harsh environments.

Photosynthetic water oxidation by Photosystem II (PSII) is a fascinating process because it sustains life on Earth and serves as a blue print for scalable synthetic catalysts required for renewable energy applications. The biophysical, computational, and structural description of this process, which started more than 50 years ago, has made tremendous progress over the past two decades, with its high-resolution crystal structures being available not only of the dark-stable state of PSII, but of all the semi-stable reaction intermediates and even some transient states. Here, we summarize the current knowledge on PSII with emphasis on the basic principles that govern the conversion of light energy to chemical energy in PSII, as well as on the illustration of the molecular structures that enable these reactions. The important remaining questions regarding the mechanism of biological water oxidation are highlighted, and one possible pathway for this fundamental reaction is described at a molecular level.

Green organisms evolve oxygen (O2) via photosynthesis and consume it by respiration. Generally, net O2 consumption only becomes dominant when photosynthesis is suppressed at night. Here, we show that green thylakoid membranes of Scots pine (Pinus sylvestris L) and Norway spruce (Picea abies) needles display strong O2 consumption even in the presence of light when extremely low temperatures coincide with high solar irradiation during early spring (ES). By employing different electron transport chain inhibitors, we show that this unusual light-induced O2 consumption occurs around photosystem (PS) I and correlates with higher abundance of flavodiiron (Flv) A protein in ES thylakoids. With P700 absorption changes, we demonstrate that electron scavenging from the acceptor-side of PSI via O2 photoreduction is a major alternative pathway in ES. This photoprotection mechanism in vascular plants indicates that conifers have developed an adaptative evolution trajectory for growing in harsh environments.

Photosynthetic water oxidation by Photosystem II (PSII) is a fascinating process because it sustains life on Earth and serves as a blue print for scalable synthetic catalysts required for renewable energy applications. The biophysical, computational, and structural description of this process, which started more than 50 years ago, has made tremendous progress over the past two decades, with its high-resolution crystal structures being available not only of the dark-stable state of PSII, but of all the semi-stable reaction intermediates and even some transient states. Here, we summarize the current knowledge on PSII with emphasis on the basic principles that govern the conversion of light energy to chemical energy in PSII, as well as on the illustration of the molecular structures that enable these reactions. The important remaining questions regarding the mechanism of biological water oxidation are highlighted, and one possible pathway for this fundamental reaction is described at a molecular level.

Green organisms evolve oxygen (O2) via photosynthesis and consume it by respiration. Generally, net O2 consumption only becomes dominant when photosynthesis is suppressed at night. Here, we show that green thylakoid membranes of Scots pine (Pinus sylvestris L) and Norway spruce (Picea abies) needles display strong O2 consumption even in the presence of light when extremely low temperatures coincide with high solar irradiation during early spring (ES). By employing different electron transport chain inhibitors, we show that this unusual light-induced O2 consumption occurs around photosystem (PS) I and correlates with higher abundance of flavodiiron (Flv) A protein in ES thylakoids. With P700 absorption changes, we demonstrate that electron scavenging from the acceptor-side of PSI via O2 photoreduction is a major alternative pathway in ES. This photoprotection mechanism in vascular plants indicates that conifers have developed an adaptative evolution trajectory for growing in harsh environments.

Photosynthetic water oxidation by Photosystem II (PSII) is a fascinating process because it sustains life on Earth and serves as a blue print for scalable synthetic catalysts required for renewable energy applications. The biophysical, computational, and structural description of this process, which started more than 50 years ago, has made tremendous progress over the past two decades, with its high-resolution crystal structures being available not only of the dark-stable state of PSII, but of all the semi-stable reaction intermediates and even some transient states. Here, we summarize the current knowledge on PSII with emphasis on the basic principles that govern the conversion of light energy to chemical energy in PSII, as well as on the illustration of the molecular structures that enable these reactions. The important remaining questions regarding the mechanism of biological water oxidation are highlighted, and one possible pathway for this fundamental reaction is described at a molecular level.

Photosynthetic water oxidation by Photosystem II (PSII) is a fascinating process because it sustains life on Earth and serves as a blue print for scalable synthetic catalysts required for renewable energy applications. The biophysical, computational, and structural description of this process, which started more than 50 years ago, has made tremendous progress over the past two decades, with its high-resolution crystal structures being available not only of the dark-stable state of PSII, but of all the semi-stable reaction intermediates and even some transient states. Here, we summarize the current knowledge on PSII with emphasis on the basic principles that govern the conversion of light energy to chemical energy in PSII, as well as on the illustration of the molecular structures that enable these reactions. The important remaining questions regarding the mechanism of biological water oxidation are highlighted, and one possible pathway for this fundamental reaction is described at a molecular level.

The regulation of photosynthetic machinery with a nonoxidative approach is a powerful but challenging strategy for the selective inhibition of bloom-forming cyanobacteria. Acetylacetone (AA) was recently found to be a target-selective cyanocide for Microcystis aeruginosa, but the cause and effect in the studied system are still unclear. By recording of the chemical fingerprints of the cells at two treatment intervals (12 and 72 h with 0.1 mM AA) with omics assays, the molecular mechanism of AA in inactivating Microcystis aeruginosa was elucidated. The results clearly reveal the effect of AA on ferredoxin and the consequent effects on the physiological and biochemical processes of Microcystis aeruginosa. In addition to its role as an electron acceptor of photosystem I, ferredoxin plays pivotal roles in the assimilation of nitrogen in cyanobacterial cells. The effect of AA on ferredoxin and on nonheme iron of photosystem II first cut off the photosynthetic electron transfer flow and then interrupted the synthesis of adenosine triphosphate (ATP) and reduced nicotinamide adenine dinucleotide phosphate (NADPH), which ultimately might affect carbon fixation and nitrogen assimilation metabolisms. The results here provide missing pieces in the current knowledge on the selective inhibition of cyanobacteria, which should shed light on the better control of harmful blooms.

Photosynthetic electron transfer comprises a series of light-induced redox reactions catalysed by multiprotein machinery in the thylakoid. These protein complexes possess cofactors susceptible to redox modifications by reactive small molecules. The gaseous radical nitric oxide (NO), a key signalling molecule in green algae and plants, has earlier been shown to bind to Photosystem (PS) II and obstruct electron transfer in plants. The effects of NO on cyanobacterial bioenergetics however, have long remained obscure. In this study, we exposed the model cyanobacterium Synechocystis sp. PCC 6803 to NO under anoxic conditions and followed changes in whole-cell fluorescence and oxidoreduction of P700 in vivo. Our results demonstrate that NO blocks photosynthetic electron transfer in cells by repressing PSII, PSI, and likely the NDH dehydrogenase-like complex 1 (NDH-1). We propose that iron‑sulfur clusters of NDH-1 complex may be affected by NO to such an extent that ferredoxin-derived electron injection to the plastoquinone pool, and thus cyclic electron transfer, may be inhibited. These findings reveal the profound effects of NO on Synechocystis cells and demonstrate the importance of controlled NO homeostasis in cyanobacteria.

We celebrate Govindjee, Professor Emeritus of Plant Biology, Biochemistry, and Biophysics, University of Illinois at Urbana-Champaign, on the occasion of his 90th birthday. He is renowned for his pioneering work in the discovery of the two-light reactions and two photosystems, PSI and PSII, leading to the Z-scheme of the electron transport chain; and for breakthrough advances in oxygenic photosynthesis. Govindjee’s publications have been cited over 26,000 times. He is an elected Fellow of the American Association of the Advancement of Science, USA; National Academy of Agriculture Science, India; and National Academy of Sciences, India; and beyond that, he has received many awards from scientific societies, most recently, the ISPR Lifetime Achievement Award, August 2022. As even today, Govindjee continues to actively contribute to the field, we highlight the major events in his colorful personal and rigorous scientific life with emphasis on the work done after his retirement, and as well, his prodigious accomplishments as teacher, editor, and science historian., Correction: Govindjee’s 90th birthday: a life dedicated to photosynthesis, Plant Physiology Reports, 2022, DOI: 10.1007/s40502-022-00699-0

On the occasion of the 90th birthday of Govindjee, Professor Emeritus of Plant Biology, Biochemistry, and Biophysics, the University of Illinois at Urbana-Champaign (UIUC), over 100 celebrants have sent felicitations and messages to thank him for mentoring, nurturing, and building the community of photosynthesis researchers belonging to four generations; and in making the scientific knowledge accessible to students and young researchers via his monumental writings and editorial contributions. Govindjee joined UIUC in September of 1956 to study as a graduate student in the laboratory of Robert Emerson. In 1961, he joined UIUC as an Assistant Professor and retired as a full professor in 1999. He is well-known for pioneering work in oxygenic photosynthesis, leading to the current Z-scheme, and for his breakthrough advances concerning light harvesting, primary charge separation, the role of bicarbonate on the two-electron gate of photosystem II, water oxidation, nonphotochemical quenching, and the use of biophysical techniques, such as prompt fluorescence, delayed fluorescence, thermoluminescence, and nuclear magnetic resonance. Today, despite his retirement, Govindjee continues to explore several important questions in the field of photosynthesis and documents the history of science. This tribute, in turn, attempts to capture scientific collaborations, as well as scholarly and personal contributions made by Govindjee to the lives of hundreds of scholars and students worldwide.

Idag tar vi växterna för givna. För många människor är de ett bakgrundsbrus, en fond. Hos oss växtintresserade får de lite mer fokus. Vi kan urskilja arterna i den gröna massan av växtlighet och vi oroar oss för deras framtid. I den här artikeln tar tre forskare med oss på en resa genom växternas historia – från första början, genom olika klimat som har gynnat utvecklingen av växtgrupper med olika egenskaper, fram till dagens gröna mångfald. En kort resumé av de senaste två årmiljarderna med fokus på växter.

On the occasion of the 90th birthday of Govindjee, Professor Emeritus of Plant Biology, Biochemistry, and Biophysics, the University of Illinois at Urbana-Champaign (UIUC), over 100 celebrants have sent felicitations and messages to thank him for mentoring, nurturing, and building the community of photosynthesis researchers belonging to four generations; and in making the scientific knowledge accessible to students and young researchers via his monumental writings and editorial contributions. Govindjee joined UIUC in September of 1956 to study as a graduate student in the laboratory of Robert Emerson. In 1961, he joined UIUC as an Assistant Professor and retired as a full professor in 1999. He is well-known for pioneering work in oxygenic photosynthesis, leading to the current Z-scheme, and for his breakthrough advances concerning light harvesting, primary charge separation, the role of bicarbonate on the two-electron gate of photosystem II, water oxidation, nonphotochemical quenching, and the use of biophysical techniques, such as prompt fluorescence, delayed fluorescence, thermoluminescence, and nuclear magnetic resonance. Today, despite his retirement, Govindjee continues to explore several important questions in the field of photosynthesis and documents the history of science. This tribute, in turn, attempts to capture scientific collaborations, as well as scholarly and personal contributions made by Govindjee to the lives of hundreds of scholars and students worldwide.

On the occasion of the 90th birthday of Govindjee, Professor Emeritus of Plant Biology, Biochemistry, and Biophysics, the University of Illinois at Urbana-Champaign (UIUC), over 100 celebrants have sent felicitations and messages to thank him for mentoring, nurturing, and building the community of photosynthesis researchers belonging to four generations; and in making the scientific knowledge accessible to students and young researchers via his monumental writings and editorial contributions. Govindjee joined UIUC in September of 1956 to study as a graduate student in the laboratory of Robert Emerson. In 1961, he joined UIUC as an Assistant Professor and retired as a full professor in 1999. He is well-known for pioneering work in oxygenic photosynthesis, leading to the current Z-scheme, and for his breakthrough advances concerning light harvesting, primary charge separation, the role of bicarbonate on the two-electron gate of photosystem II, water oxidation, nonphotochemical quenching, and the use of biophysical techniques, such as prompt fluorescence, delayed fluorescence, thermoluminescence, and nuclear magnetic resonance. Today, despite his retirement, Govindjee continues to explore several important questions in the field of photosynthesis and documents the history of science. This tribute, in turn, attempts to capture scientific collaborations, as well as scholarly and personal contributions made by Govindjee to the lives of hundreds of scholars and students worldwide.

On the occasion of the 90th birthday of Govindjee, Professor Emeritus of Plant Biology, Biochemistry, and Biophysics, the University of Illinois at Urbana-Champaign (UIUC), over 100 celebrants have sent felicitations and messages to thank him for mentoring, nurturing, and building the community of photosynthesis researchers belonging to four generations; and in making the scientific knowledge accessible to students and young researchers via his monumental writings and editorial contributions. Govindjee joined UIUC in September of 1956 to study as a graduate student in the laboratory of Robert Emerson. In 1961, he joined UIUC as an Assistant Professor and retired as a full professor in 1999. He is well-known for pioneering work in oxygenic photosynthesis, leading to the current Z-scheme, and for his breakthrough advances concerning light harvesting, primary charge separation, the role of bicarbonate on the two-electron gate of photosystem II, water oxidation, nonphotochemical quenching, and the use of biophysical techniques, such as prompt fluorescence, delayed fluorescence, thermoluminescence, and nuclear magnetic resonance. Today, despite his retirement, Govindjee continues to explore several important questions in the field of photosynthesis and documents the history of science. This tribute, in turn, attempts to capture scientific collaborations, as well as scholarly and personal contributions made by Govindjee to the lives of hundreds of scholars and students worldwide.

On the occasion of the 90th birthday of Govindjee, Professor Emeritus of Plant Biology, Biochemistry, and Biophysics, the University of Illinois at Urbana-Champaign (UIUC), over 100 celebrants have sent felicitations and messages to thank him for mentoring, nurturing, and building the community of photosynthesis researchers belonging to four generations; and in making the scientific knowledge accessible to students and young researchers via his monumental writings and editorial contributions. Govindjee joined UIUC in September of 1956 to study as a graduate student in the laboratory of Robert Emerson. In 1961, he joined UIUC as an Assistant Professor and retired as a full professor in 1999. He is well-known for pioneering work in oxygenic photosynthesis, leading to the current Z-scheme, and for his breakthrough advances concerning light harvesting, primary charge separation, the role of bicarbonate on the two-electron gate of photosystem II, water oxidation, nonphotochemical quenching, and the use of biophysical techniques, such as prompt fluorescence, delayed fluorescence, thermoluminescence, and nuclear magnetic resonance. Today, despite his retirement, Govindjee continues to explore several important questions in the field of photosynthesis and documents the history of science. This tribute, in turn, attempts to capture scientific collaborations, as well as scholarly and personal contributions made by Govindjee to the lives of hundreds of scholars and students worldwide.

Photosystem II (PSII) of plants, algae and cyanobacteria is a specialised protein complex that uses light energy to transfer electrons from water to plastoquinone, producing molecular oxygen and reduced plastoquinone. The PSII complex includes a peripheral antenna containing chlorophyll and other pigments to absorb light, a reaction centre that utilises the excitation energy transferred to it for charge separation, cofactors that stabilise the charge pair via electron transfer reactions, a Mn4CaO5 cluster that oxidises water, and a binding pocket where plastoquinone is reduced. The electrons and protons that PSII extracts from water are employed in the overall photosynthetic process for the reduction of CO2, which provides the chemical energy for most life on Earth. PSII is the only known biological source of O2 produced from water and is responsible for the molecular oxygen in the atmosphere.

After a brief background on Otto Heinrich Warburg (1883–1970), and some of his selected research, we provide highlights, in English, of three of his papers in the 1940s—unknown to many as they were not originally published in English. They are: two brief reports on Photosynthesis, with Wilhelm Lüttgens, originally published in German, in 1944: ‘Experiment on assimilation of carbonic acid’; and ‘Further experiments on carbon dioxide assimilation’. This is followed by a regular paper, originally published in Russian, in 1946: ‘The photochemical reduction of quinone in green granules’. Since the 1944 reports discussed here are very short, their translations are included in the Appendix, but that of the 1946 paper is provided in the Supplementary Material. In all three reports, Warburg provides the first evidence for and elaborates on light-driven water oxidation coupled to reduction of added benzoquinone. These largely overlooked studies of Warburg are in stark contrast to Warburg’s well-known error in assigning the origin of the photosynthetically formed dioxygen to carbonate.

The unicellular green alga Chlamydomonas reinhardtii is capable of photosynthetic H-2 production. H-2 evolution occurs under anaerobic conditions and is difficult to sustain due to 1) competition between [FeFe]-hydrogenase (H(2)ase), the key enzyme responsible for H-2 metabolism in algae, and the Calvin-Benson-Bassham (CBB) cycle for photosynthetic reductants and 2) inactivation of H(2)ase by O-2 coevolved in photosynthesis. Recently, we achieved sustainable H-2 photoproduction by shifting algae from continuous illumination to a train of short (1 s) light pulses, interrupted by longer (9 s) dark periods. This illumination regime prevents activation of the CBB cycle and redirects photosynthetic electrons to H(2)ase. Employing membrane-inlet mass spectrometry and (H2O)-O-18, we now present clear evidence that efficient H-2 photoproduction in pulseilluminated algae depends primarily on direct water biophotolysis, where water oxidation at the donor side of photosystem II (PSII) provides electrons for the reduction of protons by H(2)ase downstream of photosystem I. This occurs exclusively in the absence of CO2 fixation, while with the activation of the CBB cycle by longer (8 s) light pulses the H-2 photoproduction ceases and instead a slow overall H-2 uptake is observed. We also demonstrate that the loss of PSII activity in DCMU-treated algae or in PSII-deficient mutant cells can be partly compensated for by the indirect (PSII-independent) H-2 photoproduction pathway, but only for a short (<1 h) period. Thus, PSII activity is indispensable for a sustained process, where it is responsible for more than 92% of the final H-2 yield.

The unicellular green alga Chlamydomonas reinhardtii is capable of photosynthetic H-2 production. H-2 evolution occurs under anaerobic conditions and is difficult to sustain due to 1) competition between [FeFe]-hydrogenase (H(2)ase), the key enzyme responsible for H-2 metabolism in algae, and the Calvin-Benson-Bassham (CBB) cycle for photosynthetic reductants and 2) inactivation of H(2)ase by O-2 coevolved in photosynthesis. Recently, we achieved sustainable H-2 photoproduction by shifting algae from continuous illumination to a train of short (1 s) light pulses, interrupted by longer (9 s) dark periods. This illumination regime prevents activation of the CBB cycle and redirects photosynthetic electrons to H(2)ase. Employing membrane-inlet mass spectrometry and (H2O)-O-18, we now present clear evidence that efficient H-2 photoproduction in pulseilluminated algae depends primarily on direct water biophotolysis, where water oxidation at the donor side of photosystem II (PSII) provides electrons for the reduction of protons by H(2)ase downstream of photosystem I. This occurs exclusively in the absence of CO2 fixation, while with the activation of the CBB cycle by longer (8 s) light pulses the H-2 photoproduction ceases and instead a slow overall H-2 uptake is observed. We also demonstrate that the loss of PSII activity in DCMU-treated algae or in PSII-deficient mutant cells can be partly compensated for by the indirect (PSII-independent) H-2 photoproduction pathway, but only for a short (<1 h) period. Thus, PSII activity is indispensable for a sustained process, where it is responsible for more than 92% of the final H-2 yield.

The unicellular green alga Chlamydomonas reinhardtii is capable of photosynthetic H-2 production. H-2 evolution occurs under anaerobic conditions and is difficult to sustain due to 1) competition between [FeFe]-hydrogenase (H(2)ase), the key enzyme responsible for H-2 metabolism in algae, and the Calvin-Benson-Bassham (CBB) cycle for photosynthetic reductants and 2) inactivation of H(2)ase by O-2 coevolved in photosynthesis. Recently, we achieved sustainable H-2 photoproduction by shifting algae from continuous illumination to a train of short (1 s) light pulses, interrupted by longer (9 s) dark periods. This illumination regime prevents activation of the CBB cycle and redirects photosynthetic electrons to H(2)ase. Employing membrane-inlet mass spectrometry and (H2O)-O-18, we now present clear evidence that efficient H-2 photoproduction in pulseilluminated algae depends primarily on direct water biophotolysis, where water oxidation at the donor side of photosystem II (PSII) provides electrons for the reduction of protons by H(2)ase downstream of photosystem I. This occurs exclusively in the absence of CO2 fixation, while with the activation of the CBB cycle by longer (8 s) light pulses the H-2 photoproduction ceases and instead a slow overall H-2 uptake is observed. We also demonstrate that the loss of PSII activity in DCMU-treated algae or in PSII-deficient mutant cells can be partly compensated for by the indirect (PSII-independent) H-2 photoproduction pathway, but only for a short (<1 h) period. Thus, PSII activity is indispensable for a sustained process, where it is responsible for more than 92% of the final H-2 yield.

The effect of bicarbonate (HCO3–) on photosystem II (PSII) activity was discovered in the 1950s, but only recently have its molecular mechanisms begun to be clarified. Two chemical mechanisms have been proposed. One is for the electron-donor side, in which mobile HCO3– enhances and possibly regulates water oxidation by acting as proton acceptor, after which it dissociates into CO2 and H2O. The other is for the electron-acceptor side, in which (i) reduction of the QA quinone leads to the release of HCO3– from its binding site on the non-heme iron and (ii) the Em potential of the QA/QA•– couple increases when HCO3– dissociates. This suggested a protective/regulatory role of HCO3– as it is known that increasing the Em of QA decreases the extent of back-reaction-associated photodamage. Here we demonstrate, using plant thylakoids, that time-resolved membrane-inlet mass spectrometry together with 13C isotope labeling of HCO3– allows donor- and acceptor-side formation of CO2 by PSII to be demonstrated and distinguished, which opens the door for future studies of the importance of both mechanisms under in vivo conditions.

Knowledge of the manganese oxidation states of the oxygen-evolving Mn4CaO5 cluster in photosystem II (PSII) is crucial toward understanding the mechanism of biological water oxidation. There is a 4 decade long debate on this topic that historically originates from the observation of a multiline electron paramagnetic resonance (EPR) signal with effective total spin of S = 1/2 in the singly oxidized S2 state of this cluster. This signal implies an overall oxidation state of either Mn(III)3Mn(IV) or Mn(III)Mn(IV)3 for the S2 state. These 2 competing assignments are commonly known as “low oxidation (LO)” and “high oxidation (HO)” models of the Mn4CaO5 cluster. Recent advanced EPR and Mn K-edge X-ray spectroscopy studies converge upon the HO model. However, doubts about these assignments have been voiced, fueled especially by studies counting the number of flash-driven electron removals required for the assembly of an active Mn4CaO5 cluster starting from Mn(II) and Mn-free PSII. This process, known as photoactivation, appeared to support the LO model since the first oxygen is reported to evolve already after 7 flashes. In this study, we improved the quantum yield and sensitivity of the photoactivation experiment by employing PSII microcrystals that retained all protein subunits after complete manganese removal and by oxygen detection via a custom built thin-layer cell connected to a membrane inlet mass spectrometer. We demonstrate that 9 flashes by a nanosecond laser are required for the production of the first oxygen, which proves that the HO model provides the correct description of the Mn4CaO5 cluster’s oxidation states.

The effect of bicarbonate (HCO3–) on photosystem II (PSII) activity was discovered in the 1950s, but only recently have its molecular mechanisms begun to be clarified. Two chemical mechanisms have been proposed. One is for the electron-donor side, in which mobile HCO3– enhances and possibly regulates water oxidation by acting as proton acceptor, after which it dissociates into CO2 and H2O. The other is for the electron-acceptor side, in which (i) reduction of the QA quinone leads to the release of HCO3– from its binding site on the non-heme iron and (ii) the Em potential of the QA/QA•– couple increases when HCO3– dissociates. This suggested a protective/regulatory role of HCO3– as it is known that increasing the Em of QA decreases the extent of back-reaction-associated photodamage. Here we demonstrate, using plant thylakoids, that time-resolved membrane-inlet mass spectrometry together with 13C isotope labeling of HCO3– allows donor- and acceptor-side formation of CO2 by PSII to be demonstrated and distinguished, which opens the door for future studies of the importance of both mechanisms under in vivo conditions.

The effect of bicarbonate (HCO3–) on photosystem II (PSII) activity was discovered in the 1950s, but only recently have its molecular mechanisms begun to be clarified. Two chemical mechanisms have been proposed. One is for the electron-donor side, in which mobile HCO3– enhances and possibly regulates water oxidation by acting as proton acceptor, after which it dissociates into CO2 and H2O. The other is for the electron-acceptor side, in which (i) reduction of the QA quinone leads to the release of HCO3– from its binding site on the non-heme iron and (ii) the Em potential of the QA/QA•– couple increases when HCO3– dissociates. This suggested a protective/regulatory role of HCO3– as it is known that increasing the Em of QA decreases the extent of back-reaction-associated photodamage. Here we demonstrate, using plant thylakoids, that time-resolved membrane-inlet mass spectrometry together with 13C isotope labeling of HCO3– allows donor- and acceptor-side formation of CO2 by PSII to be demonstrated and distinguished, which opens the door for future studies of the importance of both mechanisms under in vivo conditions.

Govindjee, Professor Emeritus of Biochemistry, Biophysics and Plant Biology at the University of Illinois at Urbana-Champaign since 1999, is renowned for his pioneering work in the light reactions of photosynthesis and important accomplishments as educator, editor, historian and advocate of photosynthesis. In his honour, we review his contributions over the last twenty years, which were often achieved in collaboration with scientists working in laboratories around the world. We start with a short presentation about Govindjee’s career and continue with a section highlighting his passion and dedication to teach the younger generations of students and researchers about photosynthesis. His research work is grouped under different areas, including primary photochemistry; the relationship of chlorophyll a fluorescence to photosynthesis; short time-scale regulation processes of photosynthesis; studies toward improving photosynthesis and biomass yields; and, mathematical modelling of photosynthesis and artificial photosynthesis. He is best known for the discovery of the key role of bicarbonate on the electron acceptor side of Photosystem II. We also present his active involvement in the recognition of scientists at conferences and beyond, as well as of those who are no longer with us. He is committed to keeping alive the pioneers and discoverers in the field of photosynthesis in the collective memory of those in the field., Special issue: Biology and biotechnological applications of microalgae and photosynthetic prokaryotes: Part 2. Guest Editors: Julian Eaton-Rye, Benoit Guieysse, Michael Packer, Tina Summerfield and Susanna Wood

The unicellular green alga Chlamydomonas reinhardtii is capable of photosynthetic H-2 production. H-2 evolution occurs under anaerobic conditions and is difficult to sustain due to 1) competition between [FeFe]-hydrogenase (H(2)ase), the key enzyme responsible for H-2 metabolism in algae, and the Calvin-Benson-Bassham (CBB) cycle for photosynthetic reductants and 2) inactivation of H(2)ase by O-2 coevolved in photosynthesis. Recently, we achieved sustainable H-2 photoproduction by shifting algae from continuous illumination to a train of short (1 s) light pulses, interrupted by longer (9 s) dark periods. This illumination regime prevents activation of the CBB cycle and redirects photosynthetic electrons to H(2)ase. Employing membrane-inlet mass spectrometry and (H2O)-O-18, we now present clear evidence that efficient H-2 photoproduction in pulseilluminated algae depends primarily on direct water biophotolysis, where water oxidation at the donor side of photosystem II (PSII) provides electrons for the reduction of protons by H(2)ase downstream of photosystem I. This occurs exclusively in the absence of CO2 fixation, while with the activation of the CBB cycle by longer (8 s) light pulses the H-2 photoproduction ceases and instead a slow overall H-2 uptake is observed. We also demonstrate that the loss of PSII activity in DCMU-treated algae or in PSII-deficient mutant cells can be partly compensated for by the indirect (PSII-independent) H-2 photoproduction pathway, but only for a short (<1 h) period. Thus, PSII activity is indispensable for a sustained process, where it is responsible for more than 92% of the final H-2 yield.

The effect of bicarbonate (HCO3–) on photosystem II (PSII) activity was discovered in the 1950s, but only recently have its molecular mechanisms begun to be clarified. Two chemical mechanisms have been proposed. One is for the electron-donor side, in which mobile HCO3– enhances and possibly regulates water oxidation by acting as proton acceptor, after which it dissociates into CO2 and H2O. The other is for the electron-acceptor side, in which (i) reduction of the QA quinone leads to the release of HCO3– from its binding site on the non-heme iron and (ii) the Em potential of the QA/QA•– couple increases when HCO3– dissociates. This suggested a protective/regulatory role of HCO3– as it is known that increasing the Em of QA decreases the extent of back-reaction-associated photodamage. Here we demonstrate, using plant thylakoids, that time-resolved membrane-inlet mass spectrometry together with 13C isotope labeling of HCO3– allows donor- and acceptor-side formation of CO2 by PSII to be demonstrated and distinguished, which opens the door for future studies of the importance of both mechanisms under in vivo conditions.

The unicellular green alga Chlamydomonas reinhardtii is capable of photosynthetic H-2 production. H-2 evolution occurs under anaerobic conditions and is difficult to sustain due to 1) competition between [FeFe]-hydrogenase (H(2)ase), the key enzyme responsible for H-2 metabolism in algae, and the Calvin-Benson-Bassham (CBB) cycle for photosynthetic reductants and 2) inactivation of H(2)ase by O-2 coevolved in photosynthesis. Recently, we achieved sustainable H-2 photoproduction by shifting algae from continuous illumination to a train of short (1 s) light pulses, interrupted by longer (9 s) dark periods. This illumination regime prevents activation of the CBB cycle and redirects photosynthetic electrons to H(2)ase. Employing membrane-inlet mass spectrometry and (H2O)-O-18, we now present clear evidence that efficient H-2 photoproduction in pulseilluminated algae depends primarily on direct water biophotolysis, where water oxidation at the donor side of photosystem II (PSII) provides electrons for the reduction of protons by H(2)ase downstream of photosystem I. This occurs exclusively in the absence of CO2 fixation, while with the activation of the CBB cycle by longer (8 s) light pulses the H-2 photoproduction ceases and instead a slow overall H-2 uptake is observed. We also demonstrate that the loss of PSII activity in DCMU-treated algae or in PSII-deficient mutant cells can be partly compensated for by the indirect (PSII-independent) H-2 photoproduction pathway, but only for a short (<1 h) period. Thus, PSII activity is indispensable for a sustained process, where it is responsible for more than 92% of the final H-2 yield.

The effect of bicarbonate (HCO3–) on photosystem II (PSII) activity was discovered in the 1950s, but only recently have its molecular mechanisms begun to be clarified. Two chemical mechanisms have been proposed. One is for the electron-donor side, in which mobile HCO3– enhances and possibly regulates water oxidation by acting as proton acceptor, after which it dissociates into CO2 and H2O. The other is for the electron-acceptor side, in which (i) reduction of the QA quinone leads to the release of HCO3– from its binding site on the non-heme iron and (ii) the Em potential of the QA/QA•– couple increases when HCO3– dissociates. This suggested a protective/regulatory role of HCO3– as it is known that increasing the Em of QA decreases the extent of back-reaction-associated photodamage. Here we demonstrate, using plant thylakoids, that time-resolved membrane-inlet mass spectrometry together with 13C isotope labeling of HCO3– allows donor- and acceptor-side formation of CO2 by PSII to be demonstrated and distinguished, which opens the door for future studies of the importance of both mechanisms under in vivo conditions.

High solar flux is known to diminish photosynthetic growth rates, reducing biomass productivity and lowering disease tolerance. Photosystem II (PSII) of plants is susceptible to photodamage (also known as photoinactivation) in strong light, resulting in severe loss of water oxidation capacity and destruction of the water‐oxidizing complex (WOC). The repair of damaged PSIIs comes at a high energy cost and requires de novo biosynthesis of damaged PSII subunits, reassembly of the WOC inorganic cofactors and membrane remodeling. Employing membrane‐inlet mass spectrometry and O2‐polarography under flashing light conditions, we demonstrate that newly synthesized PSII complexes are far more susceptible to photodamage than are mature PSII complexes. We examined these ‘PSII birth defects’ in barley seedlings and plastids (etiochloroplasts and chloroplasts) isolated at various times during de‐etiolation as chloroplast development begins and matures in synchronization with thylakoid membrane biogenesis and grana membrane formation. We show that the degree of PSII photodamage decreases simultaneously with biogenesis of the PSII turnover efficiency measured by O2‐polarography, and with grana membrane stacking, as determined by electron microscopy. Our data from fluorescence, QB‐inhibitor binding, and thermoluminescence studies indicate that the decline of the high‐light susceptibility of PSII to photodamage is coincident with appearance of electron transfer capability QA− → QB during de‐etiolation. This rate depends in turn on the downstream clearing of electrons upon buildup of the complete linear electron transfer chain and the formation of stacked grana membranes capable of longer‐range energy transfer., Special Issue: SI

Inspired by the period-four oscillation in flash-induced oxygen evolution of photosystem II discovered by Joliot in 1969, Kok performed additional experiments and proposed a five-state kinetic model for photosynthetic oxygen evolution, known as Kok’s S-state clock or cycle1,2. The model comprises four (meta)stable intermediates (S0, S1, S2 and S3) and one transient S4 state, which precedes dioxygen formation occurring in a concerted reaction from two water-derived oxygens bound at an oxo-bridged tetra manganese calcium (Mn4CaO5) cluster in the oxygen-evolving complex3–7. This reaction is coupled to the two-step reduction and protonation of the mobile plastoquinone QB at the acceptor side of PSII. Here, using serial femtosecond X-ray crystallography and simultaneous X-ray emission spectroscopy with multi-flash visible laser excitation at room temperature, we visualize all (meta)stable states of Kok’s cycle as high-resolution structures (2.04–2.08 Å). In addition, we report structures of two transient states at 150 and 400 µs, revealing notable structural changes including the binding of one additional ‘water’, Ox, during the S2→S3 state transition. Our results suggest that one water ligand to calcium (W3) is directly involved in substrate delivery. The binding of the additional oxygen Ox in the S3 state between Ca and Mn1 supports O–O bond formation mechanisms involving O5 as one substrate, where Ox is either the other substrate oxygen or is perfectly positioned to refill the O5 position during O2 release. Thus, our results exclude peroxo-bond formation in the S3 state, and the nucleophilic attack of W3 onto W2 is unlikely.

Photosynthesis has been an important field of research for more than a century, but the present concerns about energy, environment and climate have greatly intensified interest in and research on this topic. Research has progressed rapidly in recent years, and this book is an interesting read for an audience who is concerned with various ways of harnessing solar energy. Our understanding of photosynthesis can now be said to have reached encyclopedic dimensions. There have been, in the past, many good books at various levels. Our book is expected to fulfill the needs of advanced undergraduate and beginning graduate students in branches of biology, biochemistry, biophysics, and bioengineering because photosynthesis is the basis of future advances in producing more food, more biomass, more fuel, and new chemicals for our expanding global human population. Further, the basics of photosynthesis are and will be used not only for the above, but in artificial photosynthesis, an important emerging field where chemists, researchers and engineers of solar energy systems will play a major role.

This chapter compares two different techniques for monitoring photosynthetic O2 production: the widespread Clark-type O2 electrode and the more sophisticated membrane inlet mass spectrometry (MIMS) technique. We describe how a simple membrane inlet for MIMS can be made out of a commercial Clark-type cell, and outline the advantages and drawbacks of the two techniques to guide researchers in deciding which method to use. Protocols and examples are given for measuring O2 evolution rates and for determining the number of chlorophyll molecules per active photosystem II reaction center.

Photosynthesis has been an important field of research for more than a century, but the present concerns about energy, environment and climate have greatly intensified interest in and research on this topic. Research has progressed rapidly in recent years, and this book is an interesting read for an audience who is concerned with various ways of harnessing solar energy. Our understanding of photosynthesis can now be said to have reached encyclopedic dimensions. There have been, in the past, many good books at various levels. Our book is expected to fulfill the needs of advanced undergraduate and beginning graduate students in branches of biology, biochemistry, biophysics, and bioengineering because photosynthesis is the basis of future advances in producing more food, more biomass, more fuel, and new chemicals for our expanding global human population. Further, the basics of photosynthesis are and will be used not only for the above, but in artificial photosynthesis, an important emerging field where chemists, researchers and engineers of solar energy systems will play a major role.

The concept of the Z-scheme of oxygenic photosynthesis is in all the textbooks. However, its evolution is not. We focus here mainly on some of the history of its biophysical aspects. We have arbitrarily divided here the 1941-2016 period into three sub-periods: (a) Origin of the concept of two light reactions: first hinted at, in 1941, by James Franck and Karl Herzfeld; described and explained, in 1945, by Eugene Rabinowitch; and a clear hypothesis, given in 1956 by Rabinowitch, of the then available cytochrome experiments: one light oxidizing it and another reducing it; (b) Experimental discovery of the two light reactions and two pigment systems and the Z-scheme of photosynthesis: Robert Emerson's discovery, in 1957, of enhancement in photosynthesis when two light beams (one in the far-red region, and the other of shorter wavelengths) are given together than when given separately; and the 1960 scheme of Robin Hill & Fay Bendall; and (c) Evolution of the many versions of the Z-Scheme: Louis Duysens and Jan Amesz's 1961 experiments on oxidation and reduction of cytochrome f by two different wavelengths of light, followed by the work of many others for more than 50 years.

Layers of amorphous manganese oxides were directly formed on the surfaces of different carbon materials by exposing the carbon to aqueous solutions of permanganate (MnO4- ) followed by sintering at 100-400 °C. During electrochemical measurements in neutral aqueous buffer, nearly all of the MnOx /C electrodes show significant oxidation currents at potentials relevant for the oxygen evolution reaction (OER). However, by combining electrolysis with product detection by using mass spectrometry, it was found that these currents were only strictly linked to water oxidation if MnOx was deposited on graphitic carbon materials (faradaic O2 yields >90 %). On the contrary, supports containing sp3 -C were found to be unsuitable as the OER is accompanied by carbon corrosion to CO2 . Thus, choosing the "right" carbon material is crucial for the preparation of stable and efficient MnOx /C anodes for water oxidation catalysis. For MnOx on graphitic substrates, current densities of >1 mA cm-2 at η=540 mV could be maintained for at least 16 h of continuous operation at pH 7 (very good values for electrodes containing only abundant elements such as C, O, and Mn) and post-operando measurements proved the integrity of both the catalyst coating and the underlying carbon at OER conditions.

Light-induced oxidation of water by photosystem II (PS II) in plants, algae and cyanobacteria has generated most of the dioxygen in the atmosphere. PS II, a membrane-bound multi-subunit pigment protein complex, couples the one-electron photochemistry at the reaction centre with the four-electron redox chemistry of water oxidation at the Mn4CaO5 cluster in the oxygen-evolving complex (OEC). Under illumination, the OEC cycles through five intermediate S-states (S0 to S4)1, in which S1 is the dark-stable state and S3 is the last semi-stable state before O–O bond formation and O2 evolution2,3. A detailed understanding of the O–O bond formation mechanism remains a challenge, and will require elucidation of both the structures of the OEC in the different S-states and the binding of the two substrate waters to the catalytic site4–6. Here we report the use of femtosecond pulses from an X-ray free electron laser (XFEL) to obtain damage-free, room temperature structures of dark-adapted (S1), two-flash illuminated (2F; S3-enriched), and ammonia-bound two-flash illuminated (2F-NH3; S3-enriched) PS II. Although the recent 1.95 Å resolution structure of PS II at cryogenic temperature using an XFEL7 provided a damage-free view of the S1 state, measurements at room temperature are required to study the structural landscape of proteins under functional conditions8,9, and also for in situ advancement of the S-states. To investigate the water-binding site(s), ammonia, a water analogue, has been used as a marker, as it binds to the Mn4CaO5 cluster in the S2 and S3 states10. Since the ammonia-bound OEC is active, the ammonia-binding Mn site is not a substrate water site10–13. This approach, together with a comparison of the native dark and 2F states, is used to discriminate between proposed O–O bond formation mechanisms.

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.

The two-helix light harvesting like (Lil) protein Lil3 belongs to the family of chlorophyll binding light harvesting proteins of photosynthetic membranes. A function in tetrapyrrol synthesis and stabilization of geranylgeraniol reductase has been shown. Lil proteins contain the chlorophyll a/b-binding motif; however, binding of chlorophyll has not been demonstrated. We find that Lil3.2 from Arabidopsis thaliana forms heterodimers with Lil3.1 and binds chlorophyll. Lil3.2 heterodimerization (25 +/- 7.8 nM) is favored relative to homodimerization (431 +/- 59 nM). Interaction of Lil3.2 with chlorophyll a (231 +/- 49 nM) suggests that heterodimerization precedes binding of chlorophyll in Arabidopsis thatiana.

In oxygenic photosynthesis, light energy is stored in the form of chemical energy by converting CO2 and water into carbohydrates.The light-driven oxidation of water that provides the electrons and protons for the subsequent CO2 fixation takes place inphotosystem II (PSII). Recent studies show that in higher plants, HCO3– increases PSII activity by acting as a mobile acceptor ofthe protons produced by PSII. In the green alga Chlamydomonas reinhardtii, a luminal carbonic anhydrase, CrCAH3, was suggested toimprove proton removal from PSII, possibly by rapid reformation of HCO3– from CO2. In this study, we investigated the interplaybetween PSII and CrCAH3 by membrane inlet mass spectrometry and x-ray crystallography. Membrane inlet mass spectrometrymeasurements showed that CrCAH3 was most active at the slightly acidic pH values prevalent in the thylakoid lumen underillumination. Two crystal structures of CrCAH3 in complex with either acetazolamide or phosphate ions were determined at 2.6- and2.7-Å resolution, respectively. CrCAH3 is a dimer at pH 4.1 that is stabilized by swapping of the N-terminal arms, a feature notpreviously observed in a-type carbonic anhydrases. The structure contains a disulfide bond, and redox titration of CrCAH3 functionwith dithiothreitol suggested a possible redox regulation of the enzyme. The stimulating effect of CrCAH3 and CO2/HCO3– on PSIIactivity was demonstrated by comparing the flash-induced oxygen evolution pattern of wild-type and CrCAH3-less PSIIpreparations. We showed that CrCAH3 has unique structural features that allow this enzyme to maximize PSII activity at lowpH and CO2 concentration.

Cyanobacteria, algae and plants oxidize water to the O2 we breathe, and consume CO2 during the synthesis of biomass. Although 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 inside 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., Included in thesis in manuscript form.

Over 40 years ago, Joliot et al. (Photochem Photobiol 10:309-329, 1969) designed and employed an elegant and highly sensitive electrochemical technique capable of measuring O2 evolved by photosystem II (PSII) in response to trains of single turn-over light flashes. The measurement and analysis of flash-induced oxygen evolution patterns (FIOPs) has since proven to be a powerful method for probing the turnover efficiency of PSII. Stemler et al. (Proc Natl Acad Sci USA 71(12):4679-4683, 1974), in Govindjee's lab, were the first to study the effect of "bicarbonate" on FIOPs by adding the competitive inhibitor acetate. Here, we extend this earlier work by performing FIOPs experiments at various, strictly controlled inorganic carbon (Ci) levels without addition of any inhibitors. For this, we placed a Joliot-type bare platinum electrode inside a N2-filled glove-box (containing 10-20 ppm CO2) and reduced the Ci concentration simply by washing the samples in Ci-depleted media. FIOPs of spinach thylakoids were recorded either at 20-times reduced levels of Ci or at ambient Ci conditions (390 ppm CO2). Numerical analysis of the FIOPs within an extended Kok model reveals that under Ci-depleted conditions the miss probability is discernibly larger (by 2-3 %) than at ambient conditions, and that the addition of 5 mM HCO3 (-) to the Ci-depleted thylakoids largely restores the original miss parameter. Since a "mild" Ci-depletion procedure was employed, we discuss our data with respect to a possible function of free or weakly bound HCO3 (-) at the water-splitting side of PSII., Special Issues on Photosynthesis Education Honoring Govindjee

Monitoring isotopic compositions of gaseous products (e.g., H2, O2, and CO2) by time-resolved isotope-ratio membrane-inlet mass spectrometry (TR-IR-MIMS) is widely used for kinetic and functional analyses in photosynthesis research. In particular, in combination with isotopic labeling, TR-MIMS became an essential and powerful research tool for the study of the mechanism of photosynthetic water-oxidation to molecular oxygen catalyzed by the water-oxidizing complex of photosystem II. Moreover, recently, the TR-MIMS and 18O-labeling approach was successfully applied for testing newly developed catalysts for artificial water-splitting and provided important insight about the mechanism and pathways of O2 formation. In this mini-review we summarize these results and provide a brief introduction into key aspects of the TR-MIMS technique and its perspectives for future studies of the enigmatic water-splitting chemistry.

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 O(2) 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 Q(A)(-) in PSII membrane fragments with similar efficiency as plastoquinone in intact cells. Our data also confirm that at high flashing rates O(2) 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 Photosynthesis., Available online 6 April 2012