The mechanism of photosynthetic water oxidation

Photosynthetie water oxidation is unique to plants and cyanobacteria, it occurs in thylakoid membranes. The components associated with this process include: a reaction center polypeptide, having a molecular weight (Mr) of 47–50 kilodaltons (kDa), containing a reaction center chlorophyll a labeled as P680, a plastoquinol(?)-electron donor Z, a primary electron acceptor pheophytin, and a quinone electron acceptor Q_A; three ‘extrinsic’ polypeptides having Mr of approximately 17 kDa, 23 kDa, and 33 kDa; and, in all likelihood, an approximately 34 kDa ‘intrinsic’ polypeptide associated with manganese (Mn) atoms. In addition, chloride and calcium ions appear to be essential components for water oxidation. Photons, absorbed by the so-called photosystem II, provide the necessary energy for the chemical oxidation-reduction at P680; the oxidized P680 (P680⁺), then, oxidizes Z, which then oxidizes the water-manganese system contained, perhaps, in a protein matrix. The oxidation of water, leading to O₂ evolution and H⁺ release, requires four such independent acts, i.e., there is a charge accumulating device (the so-called S-states). In this minireview, we have presented our current understanding of the reaction center P680, the chemical nature of Z, a possible working model for water oxidation, and the possible roles of manganese atoms, chloride ions, and the various polypeptides, mentioned above. A comparison with cytochrome c oxidase, which is involved in the opposite process of the reduction of O₂ to H₂O, is stressed. This minireview is a prelude to the several minireviews, scheduled to be published in the forthcoming issues of Photosynthesis Research, including those on photosystem II (by H.J. van Gorkom); polypeptides of the O₂-evolving system (by D.F. Ghanotakis and C.F. Yocum); and the role of chloride in O₂ evolution (by S. Izawa).     

In memory of Thomas Turpin Bannister (1930–2018)

Plants grown in the field experience sharp changes in irradiation due to shading effects caused by clouds, other leaves, etc. The excess of absorbed light energy is dissipated by a number of mechanisms including cyclic electron transport, photorespiration, and Mehler-type reactions. This protection is essential for survival but decreases photosynthetic efficiency. All phototrophs except angiosperms harbor flavodiiron proteins (Flvs) which relieve the excess of excitation energy on the photosynthetic electron transport chain by reducing oxygen directly to water. Introduction of cyanobacterial Flv1/Flv3 in tobacco chloroplasts resulted in transgenic plants that showed similar photosynthetic performance under steady-state illumination, but displayed faster recovery of various photosynthetic parameters, including electron transport and non-photochemical quenching during dark–light transitions. They also kept the electron transport chain in a more oxidized state and enhanced the proton motive force of dark-adapted leaves. The results indicate that, by acting as electron sinks during light transitions, Flvs contribute to increase photosynthesis protection and efficiency under changing environmental conditions as those found by plants in the field.     

Arginine-82 regulates the pKa of the group responsible for the light-driven proton release in bacteriorhodopsin.

In wild-type bacteriorhodopsin light-induced proton release occurs before uptake at neutral pH. In contrast, in mutants in which R82 is replaced by a neutral residue (as in R82A and R82Q), only a small fraction of the protons is released before proton uptake at neutral pH; the major fraction is released after uptake. In R82Q the relative amounts of the two types of proton release, "early" (preceding proton uptake) and "late" (following proton uptake), are pH dependent. The main conclusions are that 1) R82 is not the normal light-driven proton release group; early proton release can be observed in the R82Q mutant at higher pH values, suggesting that the proton release group has not been eliminated. 2) R82 affects the pKa of the proton release group both in the unphotolyzed state of the pigment and during the photocycle. In the wild type (in 150 mM salt) the pKa of this group decreases from approximately 9.5 in the unphotolyzed pigment to approximately 5.8 in the M intermediate, leading to early proton release at neutral pH. In the R82 mutants the respective values of pKa of the proton release group in the unphotolyzed pigment and in M are approximately 8 and 7.5 in R82Q (in 1 M salt) and approximately 8 and 6.5 in R82K (in 150 mM KCl). Thus in R82Q the pKa of the proton release group does not decrease enough in the photocycle to allow early proton release from this group at neutral pH. 3) Early proton release in R82Q can be detected as a photocurrent signal that is kinetically distinct from those photocurrents that are due to proton movements from the Schiff base to D85 during M formation and from D96 to the Schiff base during the M-->N transition. 4) In R82Q, at neutral pH, proton uptake from the medium occurs during the formation of O. The proton is released during the O-->bacteriorhodopsin transition, probably from D85 because the normal proton release group cannot deprotonate at this pH. 5) The time constant of early proton release is increased from 85 microseconds in the wild type to 1 ms in R82Q (in 150 mM salt). This can be directly attributed to the increase in the pKa of the proton release group and also explains the uncoupling of proton release from M formation. 6) In the E204Q mutant only late proton release is observed at both neutral and alkaline pH, consistent with the idea that E204 is the proton release group. The proton release is concurrent with the O-->bacteriorhodopsin transition, as in R82Q at neutral pH.     

Salt-induced alterations of the fluorescence yield and of emission spectra in Chlorella pyrenoidosa

1. The addition of salts to the suspending medium induces a decreasein the yield of chlorophyll a fluorescence in normal and DCMU-poisonedintact algal cells of Chlorella pyrenoidosa. Potassium and sodiumacetate cause a pronounced lowering of the fluorescence at relativelylow concentrations (0.01–0.1 M). MgCl₂ and KCl cause asimilar lowering of fluorescence but at much higher concentrations(0.1–0.4 M). In contrast to sodium acetate, ammonium acetatedoes not cause any significant change in the fluorescence transient.
2. Unlike the case in isolated chloroplasts, MgCl₂ decreasestheratio of short wavelength (mainly system 2) to long wavelength(mainly system 1) emission bands in both DCMU poisoned and normalcells. Since these salt-induced changes do not appear to berelated to the redox reactions of photosynthesis, the saltsmight have caused a decrease in the mutual distance betweenthe two photosystems by changing the microstructure of the chloroplastsin vivo thereby facilitating the spillover of excitation energyfrom strongly fluorescent system 2 to weakly fluorescent system1.
3. The light induced turbidity changes in intact algal cells,asmeasured by the increase in optical density at 540 nm, isreducedin the presence of these salts. However, MgCl₂ producesthegreatest reduction while Na acetate the least, even thoughbothof these salts (at the concentrations used) cause largesuppressionof the fluorescence transient. Moreover, the lightinduced turbiditychanges were, essentially irreversible.
4. Ashigh concentrations of salts increase the osmotic potentialof the bathing medium, it seems that the osmotic changes aswell as the ionic changes in the intact algal cells are responsiblefor the fluorescence quenching and changes in the mode of excitationtransfer observed in this study. In the case of low concentrationsof salts (e.g., 0.1 M Na or K acetate) the effects are predominantlyionic, and in the case of very high concentrations of MgCl₂(0.4 M), the osmotic effects play a much larger role.

(Received July 30, 1973; )     

Bicarbonate stimulation of oxygen evolution, ferricyanide reduction and photoinactivation using isolated chloroplasts

Dependence of Hill reaction (ferricyanide reduction) by isolated(broken) chloroplasts on bicarbonate ion increases with timeof illumination (upto 4 min) in HCO₃^- free reaction mixture.The stimulation caused by HCO₃^- is independent of light intensitydown to very low intensities indicating an involvement of thision in early photochemical events of photosystem II. Oxygenevolution was found to be more dependent than ferricyanide reductionon HCO₃^-. The existence of an endogenous non-oxygen evolvingelectron donor in chloroplasts is thus suggested. HCO₃^- is alsoshown to greatly increase the rate of photoinactivation duringHill reaction. (Received March 2, 1974; )     

The quantum efficiency of proton pumping by the purple membrane of Halobacterium halobium.

The quantum yield of H+ release in purple membrane (PM) sheets, and H+ uptake in phospholipid (egg phosphatidylcholine, PC) vesicles containing PM, was measured in single turnover light flashes using a pH-sensitive dye, p-nitrophenol, with rhodopsin as an actinometer. We have also calculated the ratio of H+ released per M412 formed (an unprotonated Shiff-base intermediate formed during the photocycle). In PM sheets, the quantum yield of H+ release depends on the medium. The quantum yield of M412 is independent of salt concentration. The ratio H+/M412 is approximately 1.8 M KC; and approximately 0.64 in 10 mM KCl. Direct measurements of the quantum yield of H+ give approximately 0.7 when the PM is suspended in 0.5 M KC; and 0.25 in 10 mM KCl. Using a quantum yield for M412 formation of 0.3 (Becher and Ebrey, 1977 Biophys J. 17:185.), these measurements also give a H+/M412 approximately 2 at high salt. In PM/PC vesicles, the H+/M412 is approximately 2 at all salt concentrations. The M412 decay is biphasic and the dye absorption change is monophasic. The dissipation of the proton gradient is very slow, taking on the order of seconds. Addition of nigericin (H+/K+ antiporter) drastically reduces the pH changes observed in PM/PC vesicles. This and the observation that the proton relaxation time is much longer than the photochemical cycling time suggest that the protons are pumped across the membrane and there is no contribution as a result of reversible binding and release of protons on just one side of the membrane.     

Primary Events, Energy Transfer, and Reactions in Photosynthetic Events: Lifetime of the Excited State In Vivo: II. Bacteriochlorophyll in Photosynthetic Bacteria at Room Temperature

Lifetime of the excited state (τ) of bacteriochlorophyll (BChl) in photosynthetic bacteria, measured with a mode-locked argon laser (oscillating at 488 nm; mode locked at 56 MHz) as light source, ranged from 0.3 to 2.5 nsec. These τ values are reported with a precision of ±0.1 nsec. The value of τ at high exciting light intensity (I) was two to three times that at low intensity. For young cultures of green bacterium Chloropseudomonas ethylicum, τ ranged from 0.5 (low I) to 1.0 nsec (high I); for those of the purple bacterium Rhodospirillum rubrum, from 0.4 (low I) to 1.0 nsec (high I); and for those of the BChl b-containing Rhodopseudomonas viridis, from 1.0 (low I) to 2.5 nsec (high I). These data provide information regarding the efficiencies of the photochemical process in these bacteria. Quantum yield (ø) of BChl fluorescence, calculated from ø = τ/τ₀ (where τ₀ is the intrinsic lifetime of fluorescence), ranges from 2-6% at low intensities to 6-14% at high intensities.     

Changes in Quantum Yield of Photosynthesis in the Red Alga Porphyridium cruentum Caused by Stepwise Reduction in the Intensity of Light Preferentially Absorbed by the Phycobilins

This paper describes the relation between the quantum yield of photosynthesis in the red alga Porphyridium cruentum, and the spectral composition of light, changed by filtering white light through aqueous phycobilin solutions of increasing optical density. At sufficiently high densities of the filter solution, no measurable photosynthesis can be observed, although chlorophyll a molecules are still being excited at a significant rate, as can be proved by calculations from spectral distribution curves, and is confirmed by the occurrence of a “second Emerson effect” upon addition of orange light. An interpretation of this result, based on other experiments, will be given in a subsequent paper. A modification of the opal glass technique for reducing the effect of scattering when measuring absorption, was developed in connection with this research, and also is described in the paper.