Electron transport pathways in spinach chloroplasts. Reduction of the primary acceptor of Photosystem II by reduced nicotinamide adenine dinucleotide phosphate in the dark

doi:10.1016/0005-2728(79)90101-4

Biochimica et Biophysica Acta (BBA) - Bioenergetics

Volume 547, Issue 1, 10 July 1979, Pages 127-137

https://doi.org/10.1016/0005-2728(79)90101-4 Get rights and content

Abstract

Addition of NADPH to osmotically lysed spinach chloroplasts results in a reduction of the primary acceptor (Q) of Photosystem II. This reduction of Q reaches a maximum of 50% in chloroplasts maintained under weak illumination and requires added ferredoxin and Mg²⁺. The reaction is inhibited by (i) an antibody to ferredoxin-NADP⁺ reductase (EC 1.6.7.1), (ii) treatment of chloroplasts with N-ethylmaleimide in the presence of NADPH, (iii) disulfodisalicylidenepropanediamine, (iv) antimycin, and (v) acceptors of non-cyclic electron transport. Uncouplers of phosphorylation do not affect NADPH-driven reduction of Q.

It is proposed that electron flow from NADPH to Q may occur in the dark by a pathway utilising portions of the normal cyclic and non-cyclic electron carrier sequences. The possible in vivo role for such a pathway in redox poising of cyclic electron transport and hence in controlling the ATP/NADPH supply ratio is discussed.

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Non-photochemical reduction of thylakoid photosynthetic redox carriers in vitro: Relevance to cyclic electron flow around photosystem I?
2014, Biochimica et Biophysica Acta - Bioenergetics
Citation Excerpt :
The half-time of the Fnpr was approximately 20 s (corresponding to an apparent first-order rate constant of approximately 0.035 s− 1), reaching a maximum level of fluorescence which was 15% of that observed following a saturating actinic pulse (620 nm, 5000 μmol photons m− 2 s− 1, 400 ms duration). No increase in chlorophyll fluorescence was observed when experiments were performed in the absence of either Fd or NADPH (Fig. S1), in accordance with earlier investigations [22,24]. The kinetics of the NADPH/Fd-dependent Fnpr presented in this study are similar those observed by others in spinach and A. thaliana chloroplast preparations [15,22,24,29], with a half-time of approximately 20 s, corresponding to a first order rate constant of approximately 0.035 s− 1.
Non-photochemical (dark) increases in chlorophyll a fluorescence yield associated with non-photochemical reduction of redox carriers (F_npr) have been attributed to the reduction of plastoquinone (PQ) related to cyclic electron flow (CEF) around photosystem I. In vivo, this rise in fluorescence is associated with activity of the chloroplast plastoquinone reductase (plastid NAD(P)H:plastoquinone oxidoreductase) complex. In contrast, this signal measured in isolated thylakoids has been attributed to the activity of the protein gradient regulation-5 (PGR5)/PGR5-like (PGRL1)-associated CEF pathway. Here, we report a systematic experimentation on the origin of F_npr in isolated thylakoids. Addition of NADPH and ferredoxin to isolated spinach thylakoids resulted in the reduction of the PQ pool, but neither its kinetics nor its inhibitor sensitivities matched those of F_npr. Notably, F_npr was more rapid than PQ reduction, and completely insensitive to inhibitors of the PSII Q_B site and oxygen evolving complex as well as inhibitors of the cytochrome b₆f complex. We thus conclude that F_npr in isolated thylakoids is not a result of redox equilibrium with bulk PQ. Redox titrations and fluorescence emission spectra imply that F_npr is dependent on the reduction of a low potential redox component (E_m about − 340 mV) within photosystem II (PSII), and is likely related to earlier observations of low potential variants of Q_A within a subpopulation of PSII that is directly reducible by ferredoxin. The implications of these results for our understanding of CEF and other photosynthetic processes are discussed.
Inhibitor studies on non-photochemical plastoquinone reduction and H <inf>2</inf> photoproduction in Chlamydomonas reinhardtii
2005, Biochimica et Biophysica Acta - Bioenergetics
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It has been reported that NADPH could reduce the PQ pool in thylakoid preparations via FNR, directly [52] or indirectly via Fd reduction and subsequent PQ reduction by FQR activity [53], both activities having been described as involved in cyclic electron transfer around PSI. Also, NEM [54] and DPI [36] have been reported to inhibit FNR activity in presence of NADPH. In our experiments, the affinity of the NAD(P)H-PQ oxidoreductase was higher for NADH than for NADPH, thus arguing against a major participation of the FNR, at least for the NADH-mediated flow.
In the absence of PSII, non-photochemical reduction of plastoquinones (PQs) occurs following NADH or NADPH addition in thylakoid membranes of the green alga Chlamydomonas reinhardtii. The nature of the enzyme involved in this reaction has been investigated in vitro by measuring chlorophyll fluorescence increase in anoxia and light-dependent O₂ uptake in the presence of methyl viologen. Based on the insensitivity of these reactions to rotenone, a type-I NADH dehydrogenase (NDH-1) inhibitor, and their sensitivity to flavoenzyme inhibitors and thiol blocking agents, we conclude to the involvement of a type-II NADH dehydrogenase (NDH-2) in PQ reduction. Intact Chlamydomonas cells placed in anoxia have the property to produce H₂ in the light by a Fe-hydrogenase which uses reduced ferredoxin as an electron donor. H₂ production also occurs in the absence of PSII thanks to the existence of a non-photochemical pathway of PQ reduction. From inhibitors effects, we suggest the involvement of a plastidial NDH-2 in PSII-independent H₂ production in Chlamydomonas. These results are discussed in relation to the absence of ndh genes in Chlamydomonas plastid genome and to the existence of 7 ORFs homologous to type-II NDHs in its nuclear genome.
Non-photochemical quenching of chlorophyll a fluorescence by oxidised plastoquinone: New evidences based on modulation of the redox state of the endogenous plastoquinone pool in broken spinach chloroplasts
2005, Biochimica et Biophysica Acta - Bioenergetics
Twenty-five years ago, non-photochemical quenching of chlorophyll fluorescence by oxidised plastoquinone (PQ) was proposed to be responsible for the lowering of the maximum fluorescence yield reported to occur when leaves or chloroplasts were treated in the dark with 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), an inhibitor of electron flow beyond the primary quinone electron acceptor (Q_A) of photosystem (PS) II [C. Vernotte, A.L. Etienne, J.-M. Briantais, Quenching of the system II chlorophyll fluorescence by the plastoquinone pool, Biochim. Biophys. Acta 545 (1979) 519-527]. Since then, the notion of PQ-quenching has received support but has also been put in doubt, due to inconsistent experimental findings. In the present study, the possible role of the native PQ-pool as a non-photochemical quencher was reinvestigated, employing measurements of the fast chlorophyll a fluorescence kinetics (from 50 μs to 5 s). The about 20% lowering of the maximum fluorescence yield F_M, observed in osmotically broken spinach chloroplasts treated with DCMU, was eliminated when the oxidised PQ-pool was non-photochemically reduced to PQH₂ by dark incubation of the samples in the presence of NAD(P)H, both under anaerobic and aerobic conditions. Incubation under anaerobic conditions in the absence of NAD(P)H had comparatively minor effects. In DCMU-treated samples incubated in the presence of NAD(P)H fluorescence quenching started to develop again after 20–30 ms of illumination, i.e., the time when PQH₂ starts getting reoxidised by PS I activity. NAD(P)H-dependent restoration of F_M was largely, if not completely, eliminated when the samples were briefly (5 s) pre-illuminated with red or far-red light. Addition to the incubation medium of HgCl₂ that inhibits dark reduction of PQ by NAD(P)H also abolished NAD(P)H-dependent restoration of F_M. Collectively, our results provide strong new evidence for the occurrence of PQ-quenching. The finding that DCMU alone did not affect the minimum fluorescence yield F₀ allowed us to calculate, for different redox states of the native PQ-pool, the fractional quenching at the F₀ level (Q₀) and to compare it with the fractional quenching at the F_M level (Q_M). The experimentally determined Q₀/Q_M ratios were found to be equal to the corresponding F₀/F_M ratios, demonstrating that PQ-quenching is solely exerted on the excited state of antenna chlorophylls.
PGR5 is involved in cyclic electron flow around photosystem I and is essential for photoprotection in Arabidopsis
2002, Cell
During photosynthesis, plants must control the utilization of light energy in order to avoid photoinhibition. We isolated an Arabidopsis mutant, pgr5 (proton gradient regulation), in which downregulation of photosystem II photochemistry in response to intense light was impaired. PGR5 encodes a novel thylakoid membrane protein that is involved in the transfer of electrons from ferredoxin to plastoquinone. This alternative electron transfer pathway, whose molecular identity has long been unclear, is known to function in vivo in cyclic electron flow around photosystem I. We propose that the PGR5 pathway contributes to the generation of a ΔpH that induces thermal dissipation when Calvin cycle activity is reduced. Under these conditions, the PGR5 pathway also functions to limit the overreduction of the acceptor side of photosystem I, thus preventing photosystem I photoinhibition.
Salt shock-inducible Photosystem I cyclic electron transfer in Synechocystis PCC6803 relies on binding of ferredoxin:NADP<sup>+</sup> reductase to the thylakoid membranes via its CpcD phycobilisome-linker homologous N-terminal domain
2000, Biochimica et Biophysica Acta - Bioenergetics
Citation Excerpt :
Instead, it is proposed that the succinate dehydrogenase complex contributes to the reduction of the plastoquinone b6f domain of Synechocystis in the DM4 revertants. This is the first report that demonstrates the involvement of FNR in PS I-dependent cyclic electron flow without use of inhibitors or modifying agents as was the case in other studies with chloroplasts or cyanobacteria [16,17,41,42]. Instead the work presented is based on the characterisation of the phenotype of a mutant that was genetically engineered.
Relative to ferredoxin:NADP⁺ reductase (FNR) from chloroplasts, the comparable enzyme in cyanobacteria contains an additional 9 kDa domain at its amino-terminus. The domain is homologous to the phycocyanin associated linker polypeptide CpcD of the light harvesting phycobilisome antennae. The phenotypic consequences of the genetic removal of this domain from the petH gene, which encodes FNR, have been studied in Synechocystis PCC 6803. The in frame deletion of 75 residues at the amino-terminus, rendered chloroplast length FNR enzyme with normal functionality in linear photosynthetic electron transfer. Salt shock correlated with increased abundance of petH mRNA in the wild-type and mutant alike. The truncation stopped salt stress-inducible increase of Photosystem I-dependent cyclic electron flow. Both photoacoustic determination of the storage of energy from Photosystem I specific far-red light, and the re-reduction kinetics of P700⁺, suggest lack of function of the truncated FNR in the plastoquinone–cytochrome b₆f complex reductase step of the PS I-dependent cyclic electron transfer chain. Independent gold-immunodecoration studies and analysis of FNR distribution through activity staining after native polyacrylamide gelelectrophoresis showed that association of FNR with the thylakoid membranes of Synechocystis PCC 6803 requires the presence of the extended amino-terminal domain of the enzyme. The truncated ΔpetH gene was also transformed into a NAD(P)H dehydrogenase (NDH1) deficient mutant of Synechocystis PCC 6803 (strain M55) (T. Ogawa, Proc. Natl. Acad. Sci. USA 88 (1991) 4275–4279). Phenotypic characterisation of the double mutant supported our conclusion that both the NAD(P)H dehydrogenase complex and FNR contribute independently to the quinone cytochrome b₆f reductase step in PS I-dependent cyclic electron transfer. The distribution, binding properties and function of FNR in the model cyanobacterium Synechocystis PCC 6803 will be discussed.
The role of chloroplastic NAD(P)H dehydrogenase in photoprotection
1999, FEBS Letters
After a brief exposure to supra-saturating light, leaves of a tobacco transformant, in which chloroplastic NAD(P)H dehydrogenase (NDH) was defective, showed more severe photoinhibition than the wild-type, when judged by the parameter of chlorophyll fluorescence Fv/Fm. Repeated application of supra-saturating light eventually resulted in chlorosis in the NDH-defective mutant, while the wild-type sustained less photodamage and was able to recover from it. The mechanism of the phenomena is discussed with respect to the potential role of NDH in photosynthesis.

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^∗: Present address: Glynn Research Laboratories, Bodmin, Cornwall, PL30 4AU, U.K.

^∗∗: Present address: Section of Biochemistry, Molecular and Cell Biology, Cornell University, Ithaca, NY 14853, U.S.A.

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Electron transport pathways in spinach chloroplasts. Reduction of the primary acceptor of Photosystem II by reduced nicotinamide adenine dinucleotide phosphate in the dark

Abstract

Biochim. Biophys. Acta

Biochim. Biophys. Acta

Biochim. Biophys. Acta

J. Biol. Chem.

Biochim. Biophys. Acta

Biochim. Biophys. Acta

J. Theor. Biol.

Biochim. Biophys. Acta

Biochim. Biophys. Acta

Methods Enzymol.

FEBS Lett.

Eur. J. Biochem.