Pseudocyclic Electron Flow Research Articles

Electron transport in chloroplasts: regulation and alternative pathways of electron transfer

This work represents an overview of electron transport regulation in chloroplasts as considered in the context of structure-function organization of photosynthetic apparatus in plants. A basic focus of the article is concentrated on a bifurcated oxidation of plastoquinol by the cytochrome b6f complex, which represents the rate-limiting step of electron transfer between photosystems 2 and 1. Electron transport along the chains of the noncyclic, cyclic and pseudocyclic electron flow, their relationships to generation of the trans-thylakoid difference in electrochemical potentials of protons in chloroplasts, and the pH-dependent mechanisms of regulation of the cytochrome b6f complex, are considered. Redox reactions with the participation of molecular oxygen and ascorbate, the alternative mediators of electron transport in chloroplasts, have also been discussed.

Electron Transport in Chloroplasts: Regulation and Alternative Pathways of Electron Transfer.

This work represents an overview of electron transport regulation in chloroplasts as considered in the context of structure-function organization of photosynthetic apparatus in plants. Main focus of the article is on bifurcated oxidation of plastoquinol by the cytochrome b6f complex, which represents the rate-limiting step of electron transfer between photosystems II and I. Electron transport along the chains of non-cyclic, cyclic, and pseudocyclic electron flow, their relationships to generation of the trans-thylakoid difference in electrochemical potentials of protons in chloroplasts, and pH-dependent mechanisms of regulation of the cytochrome b6f complex are considered. Redox reactions with participation of molecular oxygen and ascorbate, alternative mediators of electron transport in chloroplasts, have also been discussed.

Alternative electron transport pathways contribute to tolerance to high light stress in lichenized algae.

The photosynthetic apparatus of lichen photobionts has been well-characterized by chlorophyll fluorescence analysis (e.g., by pulse amplitude modulation [PAM]), which provides a proxy of the activity of photosystem II (PSII) and its antenna. However, such kinetics are unable to directly characterize photosystem I (PSI) activity and the associated alternative electron pathways that may be involved in photoprotection. Instead, PSI can be probed in vivo by near-infrared absorption, measured at the same time as standard chlorophyll fluorescence (e.g., using the WALZ Dual PAM). Here, we used the Dual PAM to investigate cyclic electron flow and photoprotection in a range of mostly temperate lichens sampled from shaded to more open microhabitats. Sun species displayed lower acceptor side limitation of PSI (Y[NA]) early in illumination when compared to shade species, indicative of higher flavodiiron-mediated pseudocyclic electron flow. In response to high irradiance, some lichens accumulate melanin, and Y[NA] was lower and NAD(P)H dehydrogenase (NDH-2)-type cyclic flow was higher in melanised than pale forms. Furthermore, non-photochemical quenching (NPQ) was higher and faster relaxing in shade than sun species, while all lichens displayed high rates of photosynthetic cyclic electron flow. In conclusion, our data suggest that (1) low acceptor side limitation of PSI is important for sun-exposed lichens; (2) NPQ helps shade species tolerate brief exposure to high irradiance; and (3) cyclic electron flow is a prominent feature of lichens regardless of habitat, although NDH-2-type flow is associated with high light acclimation.

Regulation of electron transport is essential for photosystem I stability and plant growth.

Photosynthetic electron transport is regulated by cyclic and pseudocyclic electron flow (CEF and PCEF) to maintain the balance between light availability and metabolic demands. CEF transfers electrons from photosystem I to the plastoquinone pool with two mechanisms, dependent either on PGR5/PGRL1 or on the type I NADH dehydrogenase-like (NDH) complex. PCEF uses electrons from photosystem I to reduce oxygen and in many groups of photosynthetic organisms, but remarkably not in angiosperms, it is catalyzed by flavodiiron proteins (FLVs). In this study, Physcomitrella patens plants depleted in PGRL1, NDH and FLVs in different combinations were generated and characterized, showing that all these mechanisms are active in this moss. Surprisingly, in contrast to flowering plants, Physcomitrella patens can cope with the simultaneous inactivation of PGR5- and NDH-dependent CEF but, when FLVs are also depleted, plants show strong growth reduction and photosynthetic activity is drastically reduced. The results demonstrate that mechanisms for modulation of photosynthetic electron transport have large functional overlap but are together indispensable to protect photosystem I from damage and they are an essential component for photosynthesis in any light regime.

Enhanced Production of D-Lactate in Cyanobacteria by Re-Routing Photosynthetic Cyclic and Pseudo-Cyclic Electron Flow.

Cyanobacteria are promising chassis strains for the photosynthetic production of platform and specialty chemicals from carbon dioxide. Their efficient light harvesting and metabolic flexibility abilities have allowed a wide range of biomolecules, such as the bioplastic polylactate precursor D-lactate, to be produced, though usually at relatively low yields. In order to increase photosynthetic electron flow towards the production of D-lactate, we have generated several strains of the marine cyanobacterium Synechococcus sp. PCC 7002 (Syn7002) with deletions in genes involved in cyclic or pseudo-cyclic electron flow around photosystem I. Using a variant of the Chlamydomonas reinhardtii D-lactate dehydrogenase (LDHSRT, engineered to efficiently utilize NADPH in vivo), we have shown that deletion of either of the two flavodiiron flv homologs (involved in pseudo-cyclic electron transport) or the Syn7002 pgr5 homolog (proposed to be a vital part of the cyclic electron transport pathway) is able to increase D-lactate production in Syn7002 strains expressing LDHSRT and the Escherichia coli LldP (lactate permease), especially at low temperature (25°C) and 0.04% (v/v) CO2, though at elevated temperatures (38°C) and/or high (1%) CO2 concentrations, the effect was less obvious. The Δpgr5 background seemed to be particularly beneficial at 25°C and 0.04% (v/v) CO2, with a nearly 7-fold increase in D-lactate accumulation in comparison to the wild-type background (≈1000 vs ≈150 mg/L) and decreased side effects in comparison to the flv deletion strains. Overall, our results show that manipulation of photosynthetic electron flow is a viable strategy to increase production of platform chemicals in cyanobacteria under ambient conditions.

Role and regulation of class-C flavodiiron proteins in photosynthetic organisms.

The regulation of photosynthesis is crucial to efficiently support the assimilation of carbon dioxide and to prevent photodamage. One key regulatory mechanism is the pseudo-cyclic electron flow (PCEF) mediated by class-C flavodiiron proteins (FLVs). These enzymes use electrons coming from Photosystem I (PSI) to reduce oxygen to water, preventing over-reduction in the acceptor side of PSI. FLVs are widely distributed among organisms performing oxygenic photosynthesis and they have been shown to be fundamental in many different conditions such as fluctuating light, sulfur deprivation and plant submersion. Moreover, since FLVs reduce oxygen they can help controlling the redox status of the cell and maintaining the microoxic environment essential for processes such as nitrogen fixation in cyanobacteria. Despite these important roles identified in various species, the genes encoding for FLV proteins have been lost in angiosperms where their activity could have been at least partially compensated by a more efficient cyclic electron flow (CEF). The present work reviews the information emerged on FLV function, analyzing recent structural data that suggest FLV could be regulated through a conformational change.

Role of cyclic and pseudo-cyclic electron transport in response to dynamic light changes in Physcomitrella patens.

Photosynthetic organisms support cell metabolism by harvesting sunlight and driving the electron transport chain at the level of thylakoid membranes. Excitation energy and electron flow in the photosynthetic apparatus is continuously modulated in response to dynamic environmental conditions. Alternative electron flow around photosystem I plays a seminal role in this regulation contributing to photoprotection by mitigating overreduction of the electron carriers. Different pathways of alternative electron flow coexist in the moss Physcomitrella patens, including cyclic electron flow mediated by the PGRL1/PGR5 complex and pseudo-cyclic electron flow mediated by the flavodiiron proteins FLV. In this work, we generated P.patens plants carrying both pgrl1 and flva knock-out mutations. A comparative analysis of the WT, pgrl1, flva, and pgrl1 flva lines suggests that cyclic and pseudo-cyclic processes have a synergic role in the regulation of photosynthetic electron transport. However, although both contribute to photosystem I protection from overreduction by modulating electron flow following changes in environmental conditions, FLV activity is particularly relevant in the first seconds after a light change whereas PGRL1 has a major role upon sustained strong illumination.

Partially dissecting the steady-state electron fluxes in Photosystem I in wild-type and pgr5 and ndh mutants of Arabidopsis.

Cyclic electron flux (CEF) around Photosystem I (PS I) is difficult to quantify. We obtained the linear electron flux (LEFO2) through both photosystems and the total electron flux through PS I (ETR1) in Arabidopsis in CO2-enriched air. ΔFlux = ETR1 – LEFO2 is an upper estimate of CEF, which consists of two components, an antimycin A-sensitive, PGR5 (proton gradient regulation 5 protein)-dependent component and an insensitive component facilitated by a chloroplastic nicotinamide adenine dinucleotide dehydrogenase-like complex (NDH). Using wild type as well as pgr5 and ndh mutants, we observed that (1) 40% of the absorbed light was partitioned to PS I; (2) at high irradiance a substantial antimycin A-sensitive CEF occurred in the wild type and the ndh mutant; (3) at low irradiance a sizable antimycin A-sensitive CEF occurred in the wild type but not in the ndh mutant, suggesting an enhancing effect of NDH in low light; and (4) in the pgr5 mutant, and the wild type and ndh mutant treated with antimycin A, a residual ΔFlux existed at high irradiance, attributable to charge recombination and/or pseudo-cyclic electron flow. Therefore, in low-light-acclimated plants exposed to high light, ΔFlux has contributions from various paths of electron flow through PS I.

Reactive oxygen intermediates produced by photosynthetic electron transport are enhanced in short-day grown plants

Leaves of tobacco plants grown in short days (8h light) generate more reactive oxygen species in the light than leaves of plants grown in long days (16h light). A two fold higher level of superoxide production was observed even in isolated thylakoids from short day plants. By using specific inhibitors of photosystem II and of the cytochrome b6f complex, the site of O2 reduction could be assigned to photosystem I. The higher rate of O2 reduction led to the formation of a higher proton gradient in thylakoids from short day plants. In the presence of an uncoupler, the differences in O2 reduction between thylakoids from short day and long day plants were abolished. The pigment content and the protein content of the major protein complexes of the photosynthetic electron transport chain were unaffected by the growth condition. Addition of NADPH, but not of NADH, to coupled thylakoids from long day plants raised the level of superoxide production to the same level as observed in thylakoids from short day plants. The hypothesis is put forward that the binding of an unknown protein permits the higher rate of pseudocyclic electron flow in thylakoids from short-day grown plants and that this putative protein plays an important role in changing the proportions of linear, cyclic and pseudocyclic electron transport in favour of pseudocyclic electron transport. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: from Natural to Articifical.

Co-regulation of dark and light reactions in three biochemical subtypes of C4 species

Regulation of light harvesting in response to changes in light intensity, CO(2) and O(2) concentration was studied in C(4) species representing three different metabolic subtypes: Sorghum bicolor (NADP-malic enzyme), Amaranthus edulis (NAD-malic enzyme), and Panicum texanum (PEP-carboxykinase). Several photosynthetic parameters were measured on the intact leaf level including CO(2) assimilation rates, O(2) evolution, photosystem II activities, thylakoid proton circuit and dissipation of excitation energy. Gross rates of O(2) evolution (J(O)₂'), measured by analysis of chlorophyll fluorescence), net rates of O(2) evolution and CO(2) assimilation responded in parallel to changes in light and CO(2) levels. The C(4) subtypes had similar energy requirements for photosynthesis since there were no significant differences in maximal quantum efficiencies for gross rates of O(2) evolution (average value = 0.072 O(2)/quanta absorbed, approximately 14 quanta per O(2) evolved). At saturating actinic light intensities, when photosynthesis was suppressed by decreasing CO(2), ATP synthase proton conductivity (g (H) (+)) responded strongly to changes in electron flow, decreasing linearly with J(O)₂', which was previously observed in C(3) plants. It is proposed that g (H) (+) is controlled at the substrate level by inorganic phosphate availability. The results suggest development of nonphotochemical quenching in C(4) plants is controlled by a decrease in g (H) (+), which causes an increase in proton motive force by restricting proton efflux from the lumen, rather than by cyclic or pseudocyclic electron flow.

Electron flow accompanying the ascorbate peroxidase cycle in maize mesophyll chloroplasts and its cooperation with linear electron flow to NADP + and cyclic electron flow in thylakoid membrane energization

Potential roles for cyclic and pseudocyclic electron flow in C4 plants are to provide ATP for the C4 cycle and, under excess light, to down-regulate PS II activity through membrane energization. Intact mesophyll chloroplasts of maize were used to evaluate forms of electron transport including the Mehler peroxidase reaction (linear electron flow to O2, formation of H2O2 which is reduced by ascorbate, and linear flow linked to reduction of oxidized ascorbate). Addition of H2O2 to isolated chloroplasts in the light in the presence of an uncoupler induced Photosystem (PS) II activity, as determined from increases in photochemical quenching of chlorophyll fluorescence (qp) and the quantum yield of PS II. H2O2 also induced dissipation of energy by thylakoid membrane energization and non-photochemical fluorescence quenching (qn), which was inhibited by addition of an uncoupler. These effects of H2O2 on qp and qn were inhibited by addition of KCN, an inhibitor of ascorbate peroxidase. The results suggest that H2O2 is reduced via ascorbate, and that the oxidized ascorbate is then reduced by linear electron flow contributing to photochemistry and thylakoid membrane energization. Evidence for function of pseudocyclic electron flow via the Mehler peroxidase reaction was obtained with only oxygen as an electron acceptor, as well as in the presence of oxaloacetate a natural electron acceptor in C4 photosynthesis. KCN decreased qp and PS II yield in the absence and presence of oxaloacetate and, in the former case, it severely reduced q_n. KCN also decreased ΔpH formation across the thylakoid membrane based on its decrease in the light-induced quenching of 9-aminoacridine fluorescence, particularly in the absence of oxaloacetate. Antimycin A, an inhibitor of cyclic electron flow, also diminished ΔpH formation. These results provide evidence for shared energization of thylakoid membranes by the Mehler peroxidase reaction, cyclic electron flow, and linear electron flow linked to the C4 pathway.

C4 Photosynthesis (The CO2-Concentrating Mechanism and Photorespiration).

Despite previous reports of no apparent photorespiration in C4 plants based on measurements of gas exchange under 2 versus 21% O2 at varying [CO2], photosynthesis in maize (Zea mays) shows a dual response to varying [O2]. The maximum rate of photosynthesis in maize is dependent on O2 (approximately 10%). This O2 dependence is not related to stomatal conductance, because measurements were made at constant intercellular CO2 concentration (Ci); it may be linked to respiration or pseudocyclic electron flow. At a given Ci, increasing [O2] above 10% inhibits both the rate of photosynthesis, measured under high light, and the maximum quantum yield, measured under limiting light ([phi]CO2). The dual effect of O2 is masked if measurements are made under only 2 versus 21% O2. The inhibition of both photosynthesis and [phi]CO2 by O2 (measured above 10% O2) with decreasing Ci increases in a very similar manner, characteristically of O2 inhibition due to photorespiration. There is a sharp increase in O2 inhibition when the Ci decreases below 50 [mu]bar of CO2. Also, increasing temperature, which favors photorespiration, causes a decrease in [phi]CO2 under limiting CO2 and 40% O2. By comparing the degree of inhibition of photosynthesis in maize with that in the C3 species wheat (Triticum aestivum) at varying Ci, the effectiveness of C4 photosynthesis in concentrating CO2 in the leaf was evaluated. Under high light, 30[deg]C, and atmospheric levels of CO2 (340 [mu]bar), where there is little inhibition of photosynthesis in maize by O2, the estimated level of CO2 around ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) in the bundle sheath compartment was 900 [mu]bar, which is about 3 times higher than the value around Rubisco in mesophyll cells of wheat. A high [CO2] is maintained in the bundle sheath compartment in maize until Ci decreases below approximately 100 [mu]bar. The results from these gas exchange measurements indicate that photorespiration occurs in maize but that the rate is low unless the intercellular [CO2] is severely limited by stress.

A cDNA clone encoding Chlamydomonas reinhardtii preferredoxin.

Fds are ubiquitous low mo1 wt proteins containing iron sulfur center(s) involved in numerous electron transfer reactions. In plants, one of the major functions of Fd is to provide the reducing power for chloroplastic NADP photoreduction. In addition, Fd is also needed for nitrite, ammonia, and sulfite assimilation as well as for pseudocyclic electron flow and fatty acid metabolism and for the light regulation of chloroplastic enzymes (Orme-Johnson, 1973). This protein is encoded by nuclear genes and is, therefore, produced as a precursor that is subsequently cleaved inside the chloroplast (Smeekens et al., 1985). We have previously purified Chlamydomonas reinhardtii Fd and determined its primary structure by direct amino acid sequencing (Schmitter et al., 1988). Based on this sequence and using the polymerase chain reaction, we have isolated a nucleotidic sequence coding for the mature portion of C. reinhardtii Fd and shown that the deduced amino acid sequence was identical with only one substitution Thr7 to Ser (Table I). In addition, we demonstrated that Escherichia coli cells were able to direct the synthesis of C. reinhardtii Fd polypeptide and to reassemble it together with an iron-sulfur center (Rogers et al., 1992). In this paper we report the sequence of a cDNA encoding C. reinhardtii preferredoxin. A Xgtll cDNA was sequenced (527 bp) and found to encode a 126-amino acid precursor with a molecular mass of 13,250 D, compared to the mature form, which contains 94 amino acids and has a molecular mass of 9,908 D. The deduced amino acid sequence was completely homologous to the sequence reported by Schmitter et al. (1988). In addition, the sequence revealed the structure of the 32-amino acid transit peptide (molecular mass 3342 D), which is as follows: MAMAMRSTFAARVGAKPAVRGARPASRMSCMA. Severa1 lines of evidence indicate that the first ATG is the initiation codon. First, it is preceded by TCA and AAA, triplets theoretically coding for Ser and Lys but rarely if ever used for nuclear genes in C. reinhardtii. Second, the flanking sequence surrounding the initiation codon (AAAAATGGC) fits well the eukaryotic translation initiation consensus (Wedel et al., 1992). Finally, the beginning of the transit peptide (MAMAM) is highly similar to the MAQM sequence reported by Wedel et al. (1992) and to the MAMAT of Hoffmann et al. (1988), and the dipeptide MA is overwhelm-

Monitoring oxygen reduction by photosystem I in whole thylakoid membranes using a photoelectrochemical cell

Thylakoid membranes isolated from spinach generate photocurrent in a photoelectrochemical cell. The activation effect of methylviologen, a photosystem I (PS I) electron acceptor known to increase the magnitude of the photocurrent generated, has been characterized. The effects of three enzymes (superoxide dismutase, glucose oxidase and catalase) on the photocurrent have been studied. All three inhibit the photocurrent to specific enzyme-dependent degrees. These results are used to demonstrate that electrons are transferred to the working electrode via an oxygen-mediated pathway similar to a Mehler-type reaction. Furthermore, the activity originates from PS I with oxygen as the acceptor and PS II as the donor. Finally, the direct relationship between the concentration of chlorophyll and the photocurrent produced is demonstrated and evidence is given to show that catalase is a non-competitive inhibitor. The photoelectrochemical cell is established as a potential new approach for monitoring pseudocyclic electron flow.

Photorespiration and Internal CO2 Accumulation in Chara corallina as Inferred from the Influence of DIC and O2 on Photosynthesis

An O(2) electrode system with a specially designed chamber for ;whorl' cell complexes of Chara corallina was used to study the combined effects of inorganic carbon and O(2) concentrations on photosynthetic O(2) evolution. At pH = 5.5 and 20% O(2), cells grown in HCO(3) (-) medium (low CO(2), pH >/= 9.0) exhibited a higher affinity for external CO(2) (K((1/2))(CO(2)) = 40 +/- 6 micromolar) than the cells grown for at least 24 hours in high-CO(2) medium (pH = 6.5), (K((1/2))(CO(2)) = 94 +/- 16 micromolar). With O(2) </= 2% in contrast, both types of cells showed a high apparent affinity (K((1/2))(CO(2)) = 50 - 52 micromolar). A Warburg effect was detectable only in the low affinity cells previously cultivated in high-CO(2) medium (pH = 6.5). The high-pH, HCO(3) (-)-grown cells, when exposed to low pH (5.5) conditions, exhibited a response indicating an ability to fix CO(2) which exceeded the CO(2) externally supplied, and the reverse situation has been observed in high-CO(2)-grown cells. At pH 8.2, the apparent photosynthetic affinity for external HCO(3) (-) (K((1/2))[HCO(3) (-)]) was 0.6 +/- 0.2 millimolar, at 20% O(2). But under low O(2) concentrations (</=2%), surprisingly, an inhibition of net O(2) evolution was elicited, which was maximal at low HCO(3) (-) concentrations. These results indicate that: (a) photorespiration occurs in this alga and can be revealed by cultivation in high-CO(2) medium, (b) Chara cells are able to accumulate CO(2) internally by means of a process apparently independent of the plasmalemma HCO(3) (-) transport system, (c) molecular oxygen appears to be required for photosynthetic utilization of exogenous HCO(3) (-): pseudocyclic electron flow, sustained by O(2) photoreduction, may produce the additional ATP needed for the HCO(3) (-) transport.

Effect of Photon Fluence Rate on Oxygen Evolution and Uptake by Chlamydomonas reinhardtii Suspensions Grown in Ambient and CO2-Enriched Air

A closed system consisting of an assimilation chamber furnished with a membrane inlet from the liquid phase connected to a mass spectrometer was used to measure O(2) evolution and uptake by Chlamydomonas reinhardtii cells grown in ambient (0.034% CO(2)) or CO(2)-enriched (5% CO(2)) air. At pH = 6.9, 28 degrees C and concentrations of dissolved inorganic carbon (DIC) saturating for photosynthesis, O(2) uptake in the light (U(o)) equaled O(2) production (E(o)) at the light compensation point (15 micromoles photons per square meter per second). E(o) and U(o) increased with increasing photon fluence rate (PFR) but were not rate saturated at 600 micromoles photons per square meter per second, while net O(2) exchange reached a saturation level near 500 micromoles photons per square meter per second which was nearly the same for both, CO(2)-grown and air-grown cells. Comparison of the U(o)/E(o) ratios between air-grown and CO(2)-grown C. reinhardtii showed higher values for air-grown cells at light intensities higher than light compensation. For both, air-grown and CO(2)-grown algae the rates of mitochondrial O(2) uptake in the dark measured immediately before and 5 minutes after illumination were much lower than U(o) at PFR saturating for net photosynthesis. We conclude that noncyclic electron flow from water to NADP(+) and pseudocyclic electron flow via photosystem I to O(2) both significantly contribute to O(2) exchange in the light. In contrast, mitochondrial respiration and photosynthetic carbon oxidation cycle are regarded as minor O(2) consuming reactions in the light in both, air-grown and CO(2)-grown cells. It is suggested that the "extra" O(2) uptake by air-grown algae provides ATP required for the energy dependent CO(2)/HCO(3) (-) concentrating mechanism known to be present in these cells.

Measurement of the ascorbate content of spinach leaf protoplasts and chloroplasts during illumination

Protoplasts prepared from spinach leaves in May and June contained substantial amounts of ascorbate (1.33±0.28 μmol mg(-1) chlorophyll), of which 30-40% was localised in the chloroplasts. During illumination, the ascorbate content was maintained at approximately the same concentration as in the dark in both protoplasts and chloroplasts, even in the absence of CO2 when pseudocyclic electron flow would be expected to be maximal. The addition of the Mehler reagent, methyl viologen, to isolated chloroplasts caused a rapid oxidation of stromal ascorbate in the light such that less than 95% of the ascorbate was oxidised after illumination for 1 min. Similarly the stromal ascorbate pool was rapidly oxidised upon the addition of H2O2. We conclude that when the intracellular ascorbate concentration is high, photosynthetically generated H2O2 can be reduced at rates comparable to its synthesis via the ascorbate-glutathione cycle. The addition of methyl viologen which catalyses rapid production of the superoxide anion, O 2 (-) or the addition of excess H2O2, overwhelms the reductive cycle and the ascorbate system becomes partially or totally oxidised.

Mechanism of Photoactivation of Electron Transport in Intact Bryopsis Chloroplasts

The mechanism of photoactivation of photosystem I electron transport was studied in intact Bryopsis corticulans chloroplasts. The evidence from chemical and photochemical studies suggests that photoactivation is a consequence of a reduction of an electron transport component, presumably ferredoxin-NADP(+) reductase. O(2) does not act as a mediator of the process but rather acts as an electron acceptor after photoactivation has occurred. We suggest that the initial function of the chloroplasts in a transition from dark to light is to initiate pseudocyclic electron flow.

Relative activities of linear and cyclic electron flows during chloroplast CO 2-fixation

Evolution of oxygen and turnover of cytochromes b-563 and ƒ were measured upon illumination of isolated intact spinach chloroplasts with a series of flashes. The flash yield of cytochrome ƒ oxidation approximated the sum of the yields of cytochrome b-563 reduction and electron transfer through Photosystem II, regardless of whether HCO − 3, 3-phosphoglycerate or O 2 served as the terminal electron acceptor. No absorbance contribution from cytochrome b-559 was discerned within the time range studied. Some pseudocyclic electron flow occurred when both HCO − 3 and 3-phosphoglycerate were omitted, and possibly also during induction of photosynthesis; however, the flash yield data suggest that O 2 is not reduced at a significant rate during steady state photosynthesis. The maximum rate of cytochrome ƒ turnover (1000 μequiv./mg chlorophyll per h) was adequate to support the highest rates of photosynthesis observed in isolated chloroplasts. These results agree with the concept that cytochrome ƒ is a component both of the linear and cyclic pathways whereas cytochrome b-563 functions only in the cyclic pathway. NH 4Cl decreased the half time of cytochrome b-563 oxidation from 11.6 to 8.2 ms and decreased the half time of cytochrome ƒ reduction from 7.2 to 2.8 ms. The cyclic and linear pathways thus seem to be jointly regulated by a transthylakoid H + gradient through a common control point on the reducing side of cytochrome ƒ. Cyclic turnover also increased during the induction phase of photosynthesis, when linear throughput is limited by the rate of utilization of NADPH. The slow rise in the P-518 transient correlated with increased cyclic activity under the above conditions. It is proposed that flexibility in the utilization of linear and cyclic pathways allows the chloroplast to generate ATP and NADPH in ratios appropriate to varying needs.

Photorespiration and Oxygen Inhibition of Photosynthesis in Chlorella pyrenoidosa

The inhibition of photosynthesis by O(2) in air-grown Chlorella pyrenoidosa was investigated using three experimental techniques (artificial leaf, aqueous method, and O(2) electrode) to measure carbon assimilation. CO(2) response curves were determined under different O(2), pH, and temperature conditions. Regardless of the experimental technique and condition, O(2) inhibition was not evident until a concentration of 50% was reached; V(max) values were reduced whereas K(m) (CO(2)) values were unaffected by the increasing O(2) concentration. The response of photosynthesis to O(2) was independent of CO(2) and HCO(3) (-) concentrations as well as temperature. Relative rates of photosynthesis showed a 4 to 5% stimulation in 2% O(2), a 12% inhibition in 50% O(2), and a 24% inhibition in 100% O(2). The inhibition by 50% O(2) was still reversible after 20 minutes exposure whereas 100% O(2) caused irreversible inhibition after only 4 minutes.The O(2) inhibition is discussed in terms of the oxygenase reaction and a Mehler reaction supporting pseudocyclic electron flow. The results are inconsistent with the proposals that photorespiration exists in these algae and that a CO(2)-concentrating mechanism suppresses the O(2) inhibition of photosynthesis.