Some microalgae, such as Chlamydomonas reinhardtii, harbor a highly flexible photosynthetic apparatus capable of using different electron acceptors, including carbon dioxide (CO2), protons, or oxygen (O-2), allowing survival in diverse habitats. During anaerobic induction of photosynthesis, molecular O-2 is produced at photosystem II, while at the photosystem I acceptor side, the reduction of protons into hydrogen (H-2) by the plastidial [FeFe]-hydrogenases primes CO2 fixation. Although the interaction between H-2 production and CO2 fixation has been studied extensively, their interplay with O-2 produced by photosynthesis has not been considered. By simultaneously measuring gas exchange and chlorophyll fluorescence, we identified an O-2 photoreduction mechanism that functions during anaerobic dark-to-light transitions and demonstrate that flavodiiron proteins (Flvs) are the major players involved in light-dependent O-2 uptake. We further show that Flv-mediated O-2 uptake is critical for the rapid induction of CO2 fixation but is not involved in the creation of the micro-oxic niches proposed previously to protect the [FeFe]-hydrogenase from O-2. By studying a mutant lacking both hydrogenases (HYDA1 and HYDA2) and both Flvs (FLVA and FLVB), we show that the induction of photosynthesis is strongly delayed in the absence of both sets of proteins. Based on these data, we propose that Flvs are involved in an important intracellular O-2 recycling process, which acts as a relay between H-2 production and CO2 fixation.
Photocatalytic water splitting over La-doped NaTaO3 (NaTaO3:La) was improved by loading with Au cocatalyst which works as H-2 evolution sites. NaTaO3:La loaded with Au by an impregnation method showed higher and more steady activity for water splitting than that by a photodeposition method. This difference in the activity for water splitting was related to the O-2 photoreduction on the loaded Au cocatalyst, which is one of the backward reactions of water splitting. The impregnated spherical Au cocatalyst suppressed the O-2 photoreduction by photogenerated electrons more efficiently than the photodeposited hemispherical Au cocatalyst, because the perimeter of Au/NaTaO3:La interface which produces activated O-2 molecules was smaller in the impregnated Au than the photodeposited Au. (C) 2013 Elsevier B.V. All rights reserved.
During oxygenic photosynthesis, the reducing power generated by light energy conversion is mainly used to reduce carbon dioxide. In bacteria and archae, flavodiiron (Flv) proteins catalyze O-2 or NO reduction, thus protecting cells against oxidative or nitrosative stress. These proteins are found in cyanobacteria, mosses, and microalgae, but have been lost in angiosperms. Here, we used chlorophyll fluorescence and oxygen exchange measurement using [O-18]-labeled O-2 and a membrane inlet mass spectrometer to characterize Chlamydomonas reinhardtii flvB insertion mutants devoid of both FlvB and FlvA proteins. We show that Flv proteins are involved in a photo-dependent electron flow to oxygen, which drives most of the photosynthetic electron flow during the induction of photosynthesis. As a consequence, the chlorophyll fluorescence patterns are strongly affected in flvB mutants during a light transient, showing a lower PSII operating yield and a slower nonphotochemical quenching induction. Photoautotrophic growth of flvB mutants was indistinguishable from the wild type under constant light, but severely impaired under fluctuating light due to PSI photo damage. Remarkably, net photosynthesis of flvB mutants was higher than in the wild type during the initial hour of a fluctuating light regime, but this advantage vanished under long-term exposure, and turned into PSI photo damage, thus explaining the marked growth retardation observed in these conditions. We conclude that the C. reinhardtii Flv participates in a Mehler-like reduction of O-2, which drives a large part of the photosynthetic electron flow during a light transient and is thus critical for growth under fluctuating light regimes.
Cyclic electron flow (CEF) around PSI regulates acceptor-side limitations and has multiple functions in the green alga, Chlamydomonas reinhardtii. Here we draw on recent and historic literature and concentrate on its role in Photosystem I (PSI) photoprotection, outlining causes and consequences of damage to PSI and CEF's role as an avoidance mechanism. We outline two functions of CEF in PSI photoprotection that are both linked to luminal acidification: firstly, its action on Photosystem II with non-photochemical quenching and photosynthetic control and secondly, its action in poising the stroma to overcome acceptor-side limitation by rebalancing NADPH and ATP ratios for carbon fixation.
The flavodiiron proteins (FDPs) Flv1 and Flv3 in cyanobacteria function in photoreduction of O-2 to H2O, without concomitant formation of reactive oxygen species, known as the Mehler-like reaction. Both Flv1 and Flv3 are essential for growth under fluctuating light (FL) intensities, providing protection for PSI. Here we compared the global transcript profiles of the wild type (WT), Delta flv1 and Delta flv1/Delta flv3 grown under constant light (GL) and FL. In the WT, FL induced the largest down-regulation in transcripts involved in carbon-concentrating mechanisms (CCMs), while those of the nitrogen assimilation pathways increased as compared with GL. Already under GL the Delta flv1/Delta flv3 double mutant demonstrated a partial down-regulation of transcripts for CCM and nitrogen metabolism, while in FL conditions the transcripts for nitrogen assimilation were strongly down-regulated. Many alterations were specific only for Delta flv1/Delta flv3, and not detected in Delta flv1, suggesting that certain transcripts are affected primarily because of the lack of flv3. By constructing the strains overproducing solely either Flv1 or Flv3, we demonstrate that the homo-oligomers of these proteins also function in acclimation of cells to FL, by catalyzing reactions with as yet unidentified components, while the presence of both Flv1 and Flv3 is a prerequisite for the Mehler-like reaction and thus the electron transfer to O-2. Considering the low expression of flv1, it is unlikely that the Flv1 homo-oligomer is present in the WT.
Flavodiiron proteins (FDPs) constitute a group of modular enzymes widespread in Bacteria, Archaea and Eukarya. Synechocystis sp. PCC 6803 has four FDPs (Flv1-4), which are essential for the photoprotection of photosynthesis. A direct comparison of light-induced O-2 reduction (Mehler-like reaction) under high (3% CO2, HC) and low (air level CO2, LC) inorganic carbon conditions demonstrated that the Flv1/Flv3 heterodimer is solely responsible for an efficient steady-state O-2 photoreduction under HC, with flv2 and flv4 expression strongly down-regulated. Conversely, under LC conditions, Flv1/Flv3 acts only as a transient electron sink, due to the competing withdrawal of electrons by the highly induced NDH-1 complex. Further, in vivo evidence is provided indicating that Flv2/Flv4 contributes to the Mehler-like reaction when naturally expressed under LC conditions, or, when artificially overexpressed under HC. The O-2 photoreduction driven by Flv2/Flv4 occurs down-stream of PSI in a coordinated manner with Flv1/Flv3 and supports slow and steady-state O-2 photoreduction.
O-2 photoreduction by photosynthetic electron transfer, the Mehler reaction , was observed in all groups of oxygenic photosynthetic organisms [2-4], but the electron transport chain mediating this reaction remains unidentified. We provide the first evidence for the involvement of A-type flavoproteins; that reduce 02 directly to water in vitro. Synechocystis sp. strain PCC 6803 mutants defective in flv1 and flv3, encoding A-type flavoproteins, failed to exhibit O-2 photoreduction but performed normal photosynthesis and respiratiom We show that the light-enhanced O-2 uptake was not due to respiration or photorespiration. After dark acclimation, photooxidation of P-700 was severely depressed in mutants Deltaflv1 and Deltaflv3 but recovered after light activation of CO2 fixation, which gives P-700 an additional electron acceptor. Inhibition of CO2 fixation prevented recovery but scarcely affected P-700 oxidation in the wild-type, where the Mehler reaction provides an alternative route for electrons. We conclude that the source of electrons for O-2 photoreduction is PSI and that the highly conserved A-type flavoproteins Flv1 and Flv3 are essential for this process in vivo. We propose that in cyanobacteria, contrary to eukaryotes, the Mehler reaction produces no reactive oxygen species and may be evolutionarily related to the response of anaerobic bacteria to O-2.
We elucidated the metabolism of methylglyoxal (MG) in chloroplasts of higher plants. Spinach chloroplasts showed MG-dependent NADPH oxidation because of aldo-keto reductase (AKR) activity. K-m for MG and V-max of AKR activity were 6.5 mM and 3.3 mu mol NADPH (mg Chl)(-1) h(-1), respectively. Addition of MG to illuminated chloroplasts induced photochemical quenching (Qp) of Chl fluorescence, indicating that MG stimulated photosynthetic electron transport (PET). Furthermore, MG enhanced the light-dependent uptake of O-2 into chloroplasts. After illumination of chloroplasts, accumulation of H2O2 was observed. K-m for MG and V-max of O-2 uptake were about 100 mu M and 200 mu mol O-2 (mg Chl)(-1) h(-1), respectively. MG-dependent O-2 uptake was inhibited by 3-(3,4-dichlorophenyl)-1, 1-dimethylurea (DCMU) and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB). Under anaerobic conditions, the Qp of Chl fluorescence was suppressed. These results indicate that MG was reduced as a Hill oxidant by the photosystem I (PSI), and that O-2 was reduced to O-2 by the reduced MG. In other words, MG produced in chloroplasts is preferentially reduced by PSI rather than through AKR. This triggers a type of oxidative stress that may be referred to as 'plant diabetes', because it ultimately originates from a common metabolite of the primary pathways of sugar anabolism and catabolism.
In cyanobacteria, photorespiratory 2-phosphoglycolate (2PG) metabolism is mediated by three different routes, including one route involving the glycine decarboxylase complex (Gcv). It has been suggested that, in addition to conversion of 2PG into non-toxic intermediates, this pathway is important for acclimation to high-light. The photoreduction of O-2 (Mehler reaction), which is mediated by two flavoproteins Flv1 and Flv3 in cyanobacteria, dissipates excess reductants under high-light by the four electron-reduction of oxygen to water. Single and double mutants defective in these processes were constructed to investigate the relation between photorespiratory 2PG-metabolism and the photoreduction of O-2 in the cyanobacterium Synechocystis sp. PCC 6803. The single mutants Delta flv1, Delta flv3, and Delta gcvT, as well as the double mutant Delta flv1/Delta gcvT, were completely segregated but not the double mutant Delta flv3/Delta gcvT, suggesting that the T-protein subunit of the Gcv (GcvT) and Flv3 proteins cooperate in an essential process. This assumption is supported by the following results: (1) The mutant Delta flv3/Delta gcvT showed a considerable longer lag phase and sometimes bleached after shifts from slow (low light, air CO2) to rapid (standard light, 5% CO2) growing conditions. (2) Photoinhibition experiments indicated a decreased ability of the mutant Delta flv3/Delta gcvT to cope with high-light. (3) Fluorescence measurements showed that the photosynthetic electron chain is reduced in this mutant. Our data suggest that the photorespiratory 2PG-metabolism and the photoreduction of O-2, particularly that catalyzed by Flv3, cooperate during acclimation to high-light stress in cyanobacteria.
The flavodiiron proteins (FDPs) are involved in the detoxification of oxidative compounds, such as nitric oxide (NO) or O-2 in Archaea and Bacteria. In cyanobacteria, the FDPs Flv1 and Flv3 are essential in the light-dependent reduction of O2 downstream of PSI. Phylogenetic analysis revealed that two genes (flvA and flvB) in the genome of Chlamydomonas reinhardtii show high homology to flv1 and flv3 genes of the cyanobacterium Synechocystis sp. PCC 6803. The physiological role of these FDPs in eukaryotic green algae is not known, but it is of a special interest since these phototrophic organisms perform oxygenic photosynthesis similar to higher plants, which do not possess FDP homologs. We have analyzed the levels of flvA and flvB transcripts in C. reinhardtii cells under various environmental conditions and showed that these genes are highly expressed under ambient CO2 levels and during the early phase of acclimation to sulfur deprivation, just before the onset of anaerobiosis and the induction of efficient H-2 photoproduction. Importantly, the increase in transcript levels of the flvA and flvB genes was also corroborated by protein levels. These results strongly suggest the involvement of FLVA and FLVB proteins in alternative electron transport.