Quinone Electron Acceptor Research Articles

Impairment of photosystem 2 activity at the level of secondary quinone electron acceptor in chloroplasts treated with cobalt, nickel and zinc ions

Impairment of photosystem 2 activity at the level of secondary quinone electron acceptor in chloroplasts treated with cobalt, nickel and zinc ions

Impairment of photosystem 2 activity at the level of secondary quinone electron acceptor in chloroplasts treated with cobalt, nickel and zinc ions

The toxicity of heavy metals on photosystem 2 photochemistry, was investigated by monitoring Hill activity, fluorescence, and thermoluminescence properties of photosystem 2 (PS 2) in pea (Pisum sativum L. cv. Bombay) chloroplasts. In Co2+‐, Ni2+‐ or Zn2+‐treated chloroplasts 2,6‐dichlorophenolindophenol‐Hill activity was markedly inhibited. Addition of hydroxylamine which donates electrons close to PS 2 reaction center did not restore the PS 2 activity. Co2+‐, Ni2+ or Zn2+ also inhibited PS 2 activity supported by hydroxylamine in tris (hydroxymethyl)aminomethane (Tris)‐inactivated chloroplasts. These observations were confirmed by fluorescence transient measurements. This implies that the metal ions inhibit either the reaction center or the components of PS 2 acceptor side. Flash‐induced thermoluminescence studies revealed that the S2Q−A charge recombination was insensitive to metal ion addition. The S2Q−B charge recombination, however, was inhibited with increase in the level of Co2+, Ni2+ or Zn2+. The observed sensitivity of S2−B charge recombination in comparison to the stability of S2Q−A recombination suggests that the metal ions inhibit at the level of secondary quinone electron acceptor. QB. We suggest that Co2+, Ni2+ or Zn2+ do not block the electron flow between the primary and secondary quinone electron acceptor, but possibly, directly modify QB site, leading to the loss of PS 2 activity.

Copper Toxicity Affects Photosystem II Electron Transport at the Secondary Quinone Acceptor, QB

The nature of Cu(2+) inhibition of photosystem II (PSII) photochemistry in pea (Pisum sativum L.) thylakoids was investigated monitoring Hill activity and light emission properties of photosystem II. In Cu(2+)-inhibited thylakoids, diphenyl carbazide addition does not relieve the loss of Hill activity. The maximum yield of fluorescence induction restored by hydroxylamine in Tris-inactivated thylakoids is markedly reduced by Cu(2+). This suggests that Cu(2+) does not act on the donor side of PSII but on the reaction center of PSII or on components beyond. Thermoluminescence and delayed luminescence studies show that charge recombination between the positively charged intermediate in water oxidation cycle (S(2)) and negatively charged primary quinone acceptor of pSII (Q(A) (-)) is largely unaffected by Cu(2+). The S(2)Q(B) (-) charge recombination, however, is drastically inhibited which parallels the loss of Hill activity. This indicates that Cu(2+) inhibits photosystem II photochemistry primarily affecting the function of the secondary quinone electron acceptor, Q(B). We suggest that Cu(2+) does not block electron flow between the primary and secondary quinone acceptor but modifies the Q(B) site in such a way that it becomes unsuitable for further photosystem II photochemistry.

Absence of a bicarbonate-depletion effect in electron transfer between quinones in chromatophores and reaction centers of Rhodobacter sphaeroides

Higher plants, algae, and cyanobacteria are known to require bicarbonate ions for electron flow from the first stable electron acceptor quinone QA to the second electron acceptor quinone QB, and to the intersystem quinone pool. It has been suggested that in Photosystem II of oxygenic photosynthesis, bicarbonate ion functions to maintain the reaction center in a proper conformation and, perhaps, to provide the protons needed to stabilize the semiquinone (QB-). In this paper, we show that bicarbonate ions do not influence the electron flow, from the quinone QA to QB and beyond, in the photosynthetic bacterium Rhodobacter sphaeroides. No measurable effect of bicarbonate depletion, obtained by competition with formate, was observed on cytochrome b-561 reduction in chromatophores; on the flash-dependent oscillation of semiquinone formation in reaction centers; on electron transfer from QA- to QB; or on either the fast or slow recovery of the oxidized primary donor (P+) which reflects the P+QA- ----PQA or the P+QB- ----PQB reaction. The lack of an observed effect in Rhodobacter sphaeroides in contrast to the effect seen in Photosystem II is suggested to be due to the amino-acid sequence differences between the reaction centers of the two systems.

ChemInform Abstract: Preparation of Novel Electron‐Accepting Quinone Methide Imines and Their Polymerizations.

AbstractThe 7,7‐dicyanoquinone methides (I) react with bis(trimethylsilyl)carbodiimide (II) in the presence of TiCl4 to give the imines (III).

The reaction of H 2S with the photosynthetic water-oxidizing complex and its lack of reaction with the primary electron acceptor in spinach

Inhibition of photosynthetic water oxidation by H 2S, a substrate analog, has been investigated using equilibrium titrations and EPR spectroscopic detection of the electron donors in spinach Photosystem II (PS II) membranes and compared to inhibition by NH 2OH. Like NH 2OH, H 2S inhibits formation of the S 2 oxidation state of the water oxidizing complex by a two-step process in the dark. Initially, reversible inhibition of S 2 occurs upon binding to a high affinity site in the dark (S 1) state at a low concentration of inhibitor (50% inhibition: 0.07 μmol H 2S/mg Chl, corresponding to about 15 H 2S/PS II). This causes no loss of steady-state O 2 evolution and can be reversed by illumination at room temperature, which causes multiple turnovers. At higher concentrations, irreversible inhibition occurs due to the cooperative release of 3 out of 4 Mn 2+/PS II using mild washing conditions, with parallel loss of O 2 evolution activity. This stoichiometry of Mn release is preserved throughout the entire concentration range of inhibition by both H 2S and NH 2OH, suggesting a common binding site for at least 3, and possibly all 4, of the Mn ions which are present in PS II. This consistent with independent earlier work showing the net release of 4 Mn/PS II using stronger washing conditions, and also with EPR spectroscopic evidence assigning the S 2 multiline signal to a cluster of 3–4 Mn ions. The concentration of H 2S which induces 50% irreversible inhibition is 17-fold greater than that required for NH 2OH. Qualitatively, the weakerinhibition by H 2S is consistent with its lower oxidation potential compared to N 2OH. However, the quantitative agreement is poor, suggesting that other environmental factors must be involved in determining their relative inhibition strenghts. Unlike NH 2OH, H 2S does not affect the structure of the primary quinone electron acceptor, Q A −-His-Fe (structure by analogy to bacterial reaction centers), as seen by formation of the normal EPR signals at g = 1.8 and g = 1.9 for the photoreduced semiquinone, instead of shifted resonance at g = 2.1 observed with NH 2OH. H 2S is therfore unable to bind to the NH 2OH site on the acceptor side of PS II.

Preparation of Novel Electron-Accepting Quinone Methide Imines and Their Polymerizations

Abstract N,7,7-Tricyanoquinone methide imine (1), N,11,11-tricyano-1,4-naphthoquinone methide imine (2), and N,15,15-tricyano-9,10-anthraquinone methide imine (3) were successfully prepared as novel electron-accepting quinone methide imine compounds. 1 is homopolymerizable with free radical and anionic initiators, while 2 and 3 are not.

Quinone exchange in the Q A binding site of Photosystem II reaction center core preparations isolated from Chlamydomonas reinhardtii

We demonstrate here that it is possible, in Photosystem II reaction centers, to extract and replace plastoquinone-9 (PQ-9), the natural primary quinone electron acceptor, Q A, by either another molecule of PQ-9 or by ubiquinone-9 (UQ-9), an isoprenolog of the quinone which acts in this role in purple photosynthetic bacterial reaction centers. The extraction of PQ-9 and the reconstitution with PQ-9 or UQ-9 were carried out simultaneously, in the presence of the detergent Triton X-100, on Photosystem II reaction center core preparations from Chlamydomonas reinhardtii. Quinone exchange is demonstrated unambiguously by the replacement of the light-induced PQ − − PQ ultraviolet difference spectra with that for UQ − − UQ. That the semiquinones of UQ-9 and PQ-9 are photogenerated in the Q A binding site is demonstrated by the following arguments: a local electrochromic band shift of pheophytin (C550), arising from the charge on Q − A, is observed in both cases. The ratio of ΔI I 325 nm , the maximum ultra-violet absorbance change arising from the semiquinone anion (UQ − and PQ −), to that of C550, is approx. 6 in the case of PQ-9 centers and 3–4 in the case of UQ-9 centers. These ratios are consistent with the relative in vitro extinction coefficients of the PQ and UQ semiquinone anions at their absorption maxima in the ultraviolet, 13 and 8.2 mM −1 · cm −1, respectively (Bensasson, R. and Land, E.J. (1973) Biochim. Biophys. Acta 325, 175–181). Δ I I 325 nm , a measure of semiquinone anion concentration, and ΔI I ( C550) relax with similar rates whether the centers contain PQ-9 or UQ-9. Reaction centers which have lost the Q A quinone are in a metastable state. Rapid reassociation of quinone restores primary charge separation. Reaction centers which do not rapidly reassociate with quinone lose irreversibly the ability to demonstrate primary photoactivity even after readdition of the extracted quinone.

The redox potential of the primary quinone Q A of bacterial photosynthesis is independent of the divalent metal ion

The function of the ferrous ion which links the primary and secondary quinone electron acceptors, Q A and Q B, via histidine ligands from the reaction center protein of the photosynthetic bacterium Rbodobacter sphaeroids (Y) has been investigated by substitution with Mn(II), Cu(II) and Zn(II), using biosynthetic incorporation of the metal ion. The midpoint potential and pH dependence for reduction of Q A in chromatophore membranes were found to be indistinguishable for all four divalent ions, and in agreement with earlier results on the R26 and Ga strains of R. sphaeroides containing only iron. We therefore conclude that the divalent ion contributes no functionally significant covalent coupling to the primary acceptor and that these metals have no differential electrostatic influence. Furthermore, inhibition of photoreduction of Q B by the herbicide ametryne was found to be the same for all four samples. Thus, these four divalent ions exhibit no differential influence on the binding properties of this herbicide in the Q B site.

Involvement of bicarbonate in the protonation of the secondary quinone electron acceptor of photosystem II via the non-haem iron of the quinone-iron acceptor complex

The mechanism of the bicarbonate effect was investigated by monitoring flash-induced pH changes. In control chloroplasts the proton yields exhibit a binary oscillation with a period of four. In CO 2-depleted chloroplasts the binary oscillation disappears and only the period four pattern remains, which can be described by proton liberation in the water-oxidizing system. It is concluded that bicarbonate is involved in the protonation of Q 2− B. The affinity of bicarbonate to its binding site is much lower in the presence of dithionite. It is suggested that bicarbonate exerts its influence through being a ligand for the non-haem iron between Q A and Q B.

Studies on the limitations to photosynthesis in leaves of the atrazine-resistant mutant ofSenecio vulgaris L.

In leaves of an atrazine-resistant mutant ofSenecio vulgaris the quantum efficiency of CO2 assimilation was reduced by 21% compared to the atrazine-susceptible wild type, and at a light level twice that required to saturate photosynthesis in the wild type the CO2 fixation rate in the mutant was decreased by 15%. In leaves at steady-state photosynthesis there was a measurable increase in the reduction state of the photosystem II (PSII) primary quinone acceptor,Q A. Although this would lead to a decreased rate of PSII electron transport and may thus explain the decrease in quantum efficiency, this cannot account for the fall in the maximum rate of CO2 fixation. The atrazine-resistant mutant showed an appreciably longer photosynthetic induction time which indicates an effect on carbon metabolism; however, the response of CO2-fixation rate to intercellular CO2 concentration revealed no differences in carboxylation efficiency. There were also no differences in the ability to perform a State 1-State 2 transition between the atrazine-resistant and susceptible biotypes and no difference in the profiles of phosphorylated thylakoid polypeptides. It is concluded that the alteration of the redox equilibrium between PSII quinone electron acceptors in the atrazine-resistant biotype limits appreciably the photosynthetic efficiency in non-saturating light. Additionally, there is a further, as yet unidentified, limitation which decreases photosynthesis in the resistant mutant under light-saturating conditions.

Multiple anion effects on photosystem II in chloroplast membranes

We investigated the activity of several anions at various sites on photosystem II, in particular those associated with the Cl(-) effect (anion binding-site I) and the HCO3 (-) effect (anion binding-site II). Chlorophyll a fluorescence changes were used to monitor partial photosystem II reactions either in the oxygen-evolving mechanism or involving endogenous quinone electron acceptors. We find that anions such as NO3 (-), HCO3 (-), HCO2 (-), F(-), NO2 (-), and acetate can, depending on conditions, bind to either anion binding-site I, anion binding-site II, or both sites simultaneously. The anions N3 (-) and Au(CN)2 (-) are exceptions. In their presence, oxygen-consumption reactions are enhanced. The results demonstrate that an exclusive site or mode of action of an anion on photosystem II cannot be determined by measuring the Hill reaction alone. Anion interactions with photosystem II are shown to be very complex and, therefore, caution is advisable in interpreting related experiments. Carbonic anhydrase associated with photosystem II was also investigated as a possible target for some anion effects. In Cl(-)-depleted thylakoids, NO3 (-), stimulated both electron transport and carbonic anhydrase activity at low concentrations, while higher concentrations inhibited both. However, carbonic anhydrase was more sensitive to inhibition by NO3 (-) than was electron flow. Possible interpretations are discussed; the electron transport and carbonic anhydrase activity appear not to be functionally linked.

Photoreduction of Q A, Q B, and cytochrome b-559 in an oxygen-evolving photosystem II preparation from the thermophilic cyanobacterium Synechococcus sp

Light-induced absorption changes in an oxygen-evolving photosystem II (PS II) preparation from the thermophilic cyanobacterium Synechococcus sp. were analyzed using continuous illumination which caused the reduction of both Q A (first stable quinone electron acceptor) and Q B (second quinone electron acceptor of photosystem II). In this photosystem II preparation in the presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) the amount of Q A was estimated to be 1 per 42 chlorophylls. In the absence of DCMU, plastoquinone (1.68 per Q A) was photoreduced to plastohydroquinone within a few seconds, indicating that Q B is reduced and protonated during this period. An electrochromic band shift centered around 685 nm was observed with and without DCMU. The extent of this band shift caused by Q B reduction per electron was about a third or half of that caused by Q A reduction. A significant amount of cytochrome b-559 (0.86 per Q A) was photoreduced. Only 60% of the photoreduction of cytochrome b-559 was inhibited by a DCMU concentration that inhibited electron transfer beyond Q B, indicating that the site of the reduction of cytochrome b-559 is located before the Q B site and possibly on the donor side of PS II.

The iron-quinone acceptor complex in Rhodospirillum rubrum chromatophores studied by EPR

A new EPR signal is reported in Rhodospirillum rubrum chromatophores. The signal is attributed to Q − BFe 2+, the semiquinone-iron complex of the secondary quinone electron acceptor, on the basis of the following observations. (1) It is induced by a single laser flash given a room temperature and is stable. (2) It is present after odd-numbered flashes and absent after even-numbered flashes when a series of flashes is given. (3) When it is already present, low-temperature illumination results in the disappearance of the signal due to formation of the Q − AFe 2+Q − B state. (4) Its formation is inhibited by the presence of orthophenanthroline at normal values of pH. The Q − BFe 2+ signal has two main features, one at g = 1.93 and the other at g = 1.82. The two features have different microwave power and temperature dependences, with the g = 1.82 signal being more difficult to saturate and requiring lower temperatures to be observable. Raising the pH leads to an increase in the g = 1.82 feature, while the g = 1.93 signal decreases in amplitude. It is suggested that the two parts of the signal may represent two EPR forms due to structural heterogeneity. The low-field feature of the Q − BFe 2+ signal shifts to lower field as the pH is raised and a p K for this change seems to occur at pH 9.4. The Q − AFe 2+ signal at g = 1.88 also shifts as the pH is increased; however, the shift is less marked than that seen for Q − BFe 2+, the shift is to higher field and the range over which it occurs is wider and depends upon the temperature of Q − AFe 2+ formation. This effect may be due to a p K on a protein group being shifted to higher pH by the presence of Q − A. ortho-Phenanthroline broadens and shifts the Q − AFe 2+ signal. The inhibition of electron transfer between Q − A and Q B by ortho-phenanthroline becomes less effective at high pH. The new Q − BFe 2+ signal is unlike other semiquinone-iron signals reported in the literature in bacteria; however, it is remarkably similar to the Q − BFe 2+ signal reported in Photosystem II.

Effects of trypsin upon EPR signals arising from components of the donor side of Photosystem II

EPR signals from components functioning on the electron donor side of Photosystem II (PS II) have been monitored in PS II membranes isolated from spinach chloroplasts after treatment with trypsin at pH 7.5 and pH 6.0. The following information has been obtained. (1) The multiline manganese signal, the g = 4.1 signal and Signal II slow are lost with trypsin treatment at pH 7.5, but not at pH 6.0. (2) At pH 7.5 the multiline S 2 signal and the g = 4.1 signal are lost with approximately the same dependency on the incubation time with trypsin. At pH 6.0 trypsin treatment is known to block electron transfer between Q A and Q B (the first and the second quinone electron acceptors, respectively) allowing only a single turnover to occur. Under these conditions both the g = 4.1 signal and the multiline signal are induced by illumination at 200 K and their amplitudes are almost the same as in untreated samples. These results are interpreted as indicating that the g = 4.1 signal arises from a side path donor or from S 2 itself rather than a carrier functioning between the S states and the reaction center as previously suggested. (3) Cytochrome b-559 is converted to its oxidized low-potential form by trypsin treatment at both values of pH. At pH 6.0 the S-state turnover still occurs indicating that the presence of reduced high-potential cytochrome b-559 is not necessary for this process.

Interaction of the high-spin Fe atom in the photosystem II reaction center with the quinones Q A and Q B in purified oxygen-evolving PS II reaction center complex and in PS II particles

EPR signals in the high-spin region were studied at 10 K in photosystem II (PS II) particles and in a purified oxygen-evolving PS II reaction center complex under oxidizing conditions. PS II particles showed EPR peaks at g = 8.0 and 5.6, confirming the recent report by Petrouleas and Diner [(1986) Biochim. Biophys. Acta 849, 264-275]. Addition of 3-(3',4'-dichlorophenyl)-1,1-dimethylurea (DCMU) or o-phenanthroline shifted the peaks to be closer to g = 6.0 depending on the medium pH. On the other hand, the PS II reaction center complex showed peaks at g = 6.1 and 7.8, and at g = 6.1 and 6.4, in the absence and presence of o-phenanthroline, respectively. All these peaks were found to be decreased by the illumination at 10 K. These results suggest that the high-spin signals are due to Q 400, Fe(III) atom interacting with the PS II primary electron acceptor quinone Q A as reported and that the Fe atom also interacts with the secondary acceptor quinone Q B. This interaction seems to induce the highly asymmetric ligand coordination of the Fe atom and to be affected by DCMU and o-phenanthroline in a somewhat different manner.

Identification of Q 400, a high-potential electron acceptor of Photosystem II, with the iron of the quinone-iron acceptor complex

Measurements of the area bounded by the variable fluorescence induction curve and the maximum fluorescence yield as a function of redox potential led I. Ikegami and S. Katoh ((1973) Plant Cell Physiol. 14, 829–836) to propose the existence of a high-potential electron acceptor, Q 400 ( E m7.8 = 360 mV), associated with Photosystem II (PS II). We have generated the oxidized form of this acceptor (Q + 400) using ferricyanide and other oxidants in thylakoid membranes isolated from a mutant of Chlamydomonas reinhardtii lacking Photosystem I and the cytochrome b 6 f complex. Q + 400 was detected by a decrease in the extent of reduction of the primary quinone electron acceptor, Q A, in a low-intensity light flash exciting PS II reaction centers only once. EPR measurements in the presence of Q + 400 indicated the presence of new signals at g = 8, 6.4 and 5.5. These disappeared upon illumination at 200 K or upon reduction with ascorbate. Mössbauer absorption attributed to the Fe 2+ of the Q A-Fe 2+ acceptor complex of PS II disappeared upon addition of ferricyanide due to the formation of Fe 3+. The Fe 2+ signal was restored by subsequent addition of ascorbate. All of these spectroscopic signals show similar pH-dependent ( n = 1) midpoint potentials (approx. −60 mV / pH unit) and an E m7.5 = 370 mV. We assign the EPR signals to the Fe 3+ state of the quinone-iron acceptor. Electron transfer to the Fe 3+ is responsible for the decrease in Q A reduction upon single-hit flash excitation. The properties of the Fe 3+ Fe 2+ redox couple are consistent with those of Q + 400 Q 400 and we conclude that the iron of the Q A-Fe acceptor complex is responsible for this species.

Studies on thylakoid phosphorylation and noncyclic electron transport

The effect of thylakoid phosphorylation on noncyclic electron transport in spinach chloroplasts was investigated by measuring both the reduction of nicotinamide adenine dinucleotide phosphate (NADP) and the steady-state redox level of the primary electron acceptor quinone of photosystem II (Q) during electron flow to NADP. These data are compared with the theoretical predictions for an electron transport model which relates both the redox levels of Q and the photosystem II optical cross section to the overall velocity of noncyclic electron flow. It is demonstrated that (1) transfer of 15–20% of the photosystem II antenna to photosystem I may stimulate electron flow to NADP only if Q is less than 60–70% oxidized (this condition exists with our thylakoids, even at extremely low absorption fluxes, when the illumination is not specifically enriched in photosystem I absorbed wavelengths); (2) in phosphorylated thylakoids the steady-state redox level of Q is substantially shifted to a more oxidized one (measurements of this parameter using light of different wavelengths quantitatively support the idea that thylakoid phosphorylation leads to increased photosystem I and decreased photosystem II cross sections); (3) thylakoid phosphorylation leads to stimulated noncyclic electron flow to NADP only when the increased photosystem I antenna is able to bring about large increases in the steady-state level of oxidized Q.

Excitation trapping and primary charge stabilization in Rhodopseudomonas viridis cells, measured electrically with picosecond resolution

The transmembrane primary charge separation in the photosynthetic bacterium Rhodopseudomonas viridis was monitored by electric measurements of the light-gradient type [Trissl, H. W. & Kunze, U. (1985) Biochim. Biophys. Acta 806, 136-144]. Excitation of whole cells with 30-ps laser pulses at either 532 nm or 1064 nm gave rise to a biphasic increase of the photovoltage. The fast phase, contributing about 50% of the total, rose with an exponential time constant </=40 ps and was independent of the redox state of the quinone electron acceptor. It is assigned to the migration of the excitation energy in the antenna and its subsequent trapping by the reaction center, monitored by the ultrafast charge separation between the primary electron donor and the bacteriopheophytin intermediary acceptor. The slower phase (125 +/- 50 ps) only occurred when the quinone was oxidized and disappeared when it was reduced (either chemically or photochemically). It is assigned to the forward electron transfer from the bacteriopheophytin to the quinone. The relative amplitudes of these two electrogenic steps demonstrate that the bacteriopheophytin intermediary acceptor is located halfway between the primary donor and the quinone.

Acclimation of Tomato Leaves to Changes in Light Intensity; Effects on the Function of the Thylakoid Membrane

Davies, E. C, Chow, W. S., Le Fay, J. M. and Jordan, B. R. 1986. Acclimation of tomato leaves to changes in light intensity: effects on the function of the thylakoid membrane.—J. exp. Bot. 37: 211-220. When young tomato plants grown in high light (400 /anol quanta m 2 c ' low light (100/unol quanta m~ s~' the rate of dark re-oxidation of Q~, the first quinone electron acceptor of photosystem II, was decreased within 1-2 d. In contrast, the amount of coupling factor CF,, assayed by its ATPase activity, decreased more gradually over several days. The total chlorophyll content per unit leaf area remained relatively constant, although the chlorophyll a/chlorophyll b ratio declined. When young tomato plants grown in low light were transferred to high light, the ATPase activity of isolated thylakoids increased markedly within 1 d of transfer. This increase occurred more rapidly than changes in chlorophyll content per leaf area. In addition, in vivo chlorophyll fluorescence induction curves indicate that forward electron transfer from Q occurred more readily. The functional implications of these changes are discussed. Key words—Tomato, leaves, light intensity, thylakoid membrane. Correspondence to: Glasshouse Crops Research Institute, Worthing Road, Littlehampton, West Sussex BN17 6LP, U.K.