The nuclear-encoded chloroplast NADP-dependent malate dehydrogenase (NADP-MDH) is a key enzyme controlling the malate valve, to allow the indirect export of reducing equivalents. Arabidopsis thaliana (L.) Heynh. T-DNA insertion mutants of NADP-MDH were used to assess the role of the light-activated NADP-MDH in a typical C3 plant. Surprisingly, even when exposed to high-light conditions in short days, nadp-mdh knockout mutants were phenotypically indistinguishable from the wild type. The photosynthetic performance and typical antioxidative systems, such as the Beck–Halliwell–Asada pathway, were barely affected in the mutants in response to high-light treatment. The reactive oxygen species levels remained low, indicating the apparent absence of oxidative stress, in the mutants. Further analysis revealed a novel combination of compensatory mechanisms in order to maintain redox homeostasis in the nadp-mdh plants under high-light conditions, particularly an increase in the NTRC/2-Cys peroxiredoxin (Prx) system in chloroplasts. There were indications of adjustments in extra-chloroplastic components of photorespiration and proline levels, which all could dissipate excess reducing equivalents, sustain photosynthesis, and prevent photoinhibition in nadp-mdh knockout plants. Such metabolic flexibility suggests that the malate valve acts in concert with other NADPH-consuming reactions to maintain a balanced redox state during photosynthesis under high-light stress in wild-type plants.

In higher plants, [2Fe-2S] ferredoxin (Fd) proteins are the unique electron acceptors from photosystem I (PSI).. Whereas FdC1 was capable of electron transfer with FNR, redox potentiometry showed that it had a more positive redox potential than photosynthetic Fds by around 220 mV. These results indicate that FdC1 electron donation to FNR is prevented because it is thermodynamically unfavorable. Based on our data, we speculate that FdC1 has a specific function in conditions of acceptor limitation at PSI, and channels electrons away from NADP ؉ photoreduction.Ferredoxins (Fds) 3 are small soluble electron carrier proteins. In the final reaction of photosynthetic electron transfer (PET), photosystem I (PSI) donates electrons to Fd (1), which acts as the soluble electron donor to various acceptors in the chloroplast stroma and can also return electrons to the thylakoid in cyclic electron flow (CET) (2). The electron cascade to supply carbon fixation requires photoreduction of NADP ϩ by Fd, catalyzed by Fd-NADP(H) oxidoreductase (FNR) (3). Many other plastid enzymes accept electrons directly from Fd for metabolic processes. These include, but are not limited to, nitrite reductase and sulfite reductase, which are essential for assimilation of inorganic nitrogen and sulfur, respectively, and Fd-dependent glutamine oxoglutarate aminotransferase and fatty acid desaturase, which catalyze key steps in amino acid and fatty acid metabolism, respectively (4). In addition, Fd donation to thioredoxin via the Fd:thioredoxin reductase translates the redox state of the electron transfer chain into a regulatory signal controlling the activity of many enzymes (5). Fds are also capable of accepting electrons from NADPH via FNR, in a reversal of the photosynthetic reaction (6), allowing electron donation from reduced Fd to different acceptors under non-photosynthetic conditions. Most higher plants studied possess genes for several different Fd isoproteins (7-9). There is always an isoprotein that is more abundant in non-photosynthetic tissues and has higher affinity than photosynthetic and PetF-type Fds for FNR in the non-photosynthetic (often called "root") cascade (9, 10), where electrons are transferred from NADPH to Fd. In all plants for which we possess significant EST and cDNA information at least 2 separate photosynthetic isoproteins have been identified (7,8). In the C4-plant maize, different functions have been identified for two of the leaf-type Fds (11). There is a higher demand for ATP (which is disproportionately produced in CET) in the bundle sheath cells of NADP ϩ malic enzyme type C4 plants, and maize FdI and FdII are differentially expressed in mesophyll and bundle sheath cells, respectively (12). FdII has decreased affinity for FNR (13) and demonstrates a higher activity in CET around the photosystems, whereas FdI drives linear electron flow (11). In C3 plants, this spatial distribution is not observed, but duplicate photosynthetic Fds are still present, and there is some evi-* This work was supported by Deutsche For...

At the end of the linear photosynthetic electron transfer (PET) chain, the small soluble protein ferredoxin (Fd) transfers electrons to Fd:NADP(H) oxidoreductase (FNR), which can then reduce NADP+ to support C assimilation. In addition to this linear electron flow (LEF), Fd is also thought to mediate electron flow back to the membrane complexes by different cyclic electron flow (CEF) pathways: either antimycin A sensitive, NAD(P)H complex dependent, or through FNR located at the cytochrome b6f complex. Both Fd and FNR are present in higher plant genomes as multiple gene copies, and it is now known that specific Fd iso-proteins can promote CEF. In addition, FNR iso-proteins vary in their ability to dynamically interact with thylakoid membrane complexes, and it has been suggested that this may also play a role in CEF. We will highlight work on the different Fd-isoproteins and FNR-membrane association found in the bundle sheath (BSC) and mesophyll (MC) cell chloroplasts of the C4 plant maize. These two cell types perform predominantly CEF and LEF, and the properties and activities of Fd and FNR in the BSC and MC are therefore specialized for CEF and LEF respectively. A diversity of Fd isoproteins and dynamic FNR location has also been recorded in C3 plants, algae and cyanobacteria. This indicates that the principles learned from the extreme electron transport situations in the BSC and MC of maize might be usefully applied to understanding the dynamic transition between these states in other systems.

SignificanceIron limits the growth of photosynthetic organisms, especially in marine environments. Understanding the response of photosynthetic organisms to changing iron concentrations is therefore important for agriculture and biotechnology. We have identified a protein that is essential for the correct response to changing iron concentrations in photosynthetic bacteria (cyanobacteria). This protein was previously annotated as an electron transfer component of photosynthesis, called Fed2, and contains an iron−sulfur cluster. We tested Fed2, and found that it cannot act in photosynthetic electron transport. The corresponding gene is essential, and is highly conserved between cyanobacteria, algae, and higher plants. By specifically perturbing its function, we could show that it is essential for the low-iron response at the posttranscriptional level.

In linear photosynthetic electron transport, ferredoxin:NADP(H) oxidoreductase (FNR) transfers electrons from ferredoxin (Fd) to NADP Both NADPH and reduced Fd (Fd) are required for reductive assimilation and light/dark activation/deactivation of enzymes. FNR is therefore a hub, connecting photosynthetic electron transport to chloroplast redox metabolism. A correlation between FNR content and tolerance to oxidative stress is well established, although the precise mechanism remains unclear. We investigated the impact of altered FNR content and localization on electron transport and superoxide radical evolution in isolated thylakoids, and probed resulting changes in redox homeostasis, expression of oxidative stress markers, and tolerance to high light in planta. Our data indicate that the ratio of Fd to FNR is critical, with either too much or too little FNR potentially leading to increased superoxide production, and perception of oxidative stress at the level of gene transcription. In FNR overexpressing plants, which show more NADP(H) and glutathione pools, improved tolerance to high-light stress indicates that disturbance of chloroplast redox poise and increased free radical generation may help "prime" the plant and induce protective mechanisms. In fnr1 knock-outs, the NADP(H) and glutathione pools are more oxidized relative to the wild type, and the photoprotective effect is absent despite perception of oxidative stress at the level of gene transcription.

In Bacteria and Archaea, high-affinity potassium uptake is mediated by the ATP-driven KdpFABC complex. On the basis of the biochemical properties of the ATP-hydrolyzing subunit KdpB, the transport complex is classified as type IA P-type ATPase. However, the KdpA subunit, which promotes K + transport, clearly resembles a potassium channel, such that the KdpFABC complex represents a chimera of ion pumps and ion channels. In the present study, we demonstrate that the blending of these two groups of transporters in KdpFABC also entails a nucleotide-binding mechanism in which the KdpC subunit acts as a catalytic chaperone. This mechanism is found neither in P-type ATPases nor in ion channels, although parallels are found in ABC transporters. In the latter, the ATP nucleotide is coordinated by the LSGGQ signature motif via double hydrogen bonds at a conserved glutamine residue, which is also present in KdpC. High-affinity nucleotide binding to the Kdp-FABC complex was dependent on the presence of this conserved glutamine residue in KdpC. In addition, both ATP binding to KdpC and ATP hydrolysis activity of KdpFABC were sensitive to the accessibility, presence or absence of the hydroxyl groups at the ribose moiety of the nucleotide. Furthermore, the KdpC subunit was shown to interact with the nucleotide-binding loop of KdpB in an ATP-dependent manner around the ATP-binding pocket, thereby increasing the ATP-binding affinity by the formation of a transient KdpB ⁄ KdpC ⁄ ATP ternary complex. Structured digital abstractl kdpBN and kdpC bind by isothermal titration calorimetry (View interaction) l kdpBN and kdpC bind by nuclear magnetic resonance (View interaction)

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