Change in electron and spin density upon electron transfer to haem

Haems are the cofactors of cytochromes and important catalysts of biological electron transfer. They are composed of a planar porphyrin structure with iron coordinated at the centre. It is known from spectroscopy that ferric low-spin haem has one unpaired electron at the iron, and that this spin is paired as the haem receives an electron upon reduction (I. Bertini, C. Luchinat, NMR of Paramagnetic Molecules in Biological Systems, Benjamin/Cummins Publ. Co., Menlo Park, CA, 1986, pp. 165-170; H.M. Goff, in: A.B.P. Lever, H.B. Gray (Eds.), Iron Porphyrins, Part I, Addison-Wesley Publ. Co., Reading, MA, 1983, pp. 237-281; G. Palmer, in: A.B.P. Lever, H.B. Gray (Eds.), Iron Porphyrins, Part II, Addison-Wesley Publ. Co., Reading, MA, 1983, pp. 43-88). Here we show by quantum chemical calculations on a haem a model that upon reduction the spin pairing at the iron is accompanied by effective delocalisation of electrons from the iron towards the periphery of the porphyrin ring, including its substituents. The change of charge of the iron atom is only approx. 0.1 electrons, despite the unit difference in formal oxidation state. Extensive charge delocalisation on reduction is important in order for the haem to be accommodated in the low dielectric of a protein, and may have impact on the distance dependence of the rates of electron transfer. The lost individuality of the electron added to the haem on reduction is another example of the importance of quantum mechanical effects in biological systems.     

Comparison of the effect of excessive light on chlorophyll fluorescence (77K) and photon yield of O2 evolution in leaves of higher plants

High-light treatments (1750–2000 mol photons m^–2 · s^–1) of leaves from a number of higher-plant species invariably resulted in quenching of the maximum 77K chlorophyll fluorescence at both 692 and 734 nm (F _M, 692 and F _M, 734). The response of instantaneous fluorescence at 692 nm (F _O, 692) was complex. In leaves of some species F _O, 692 increased dramatically in others it was quenched, and in others yet it showed no marked, consistent change. Regardless of the response of F _O, 692 an apparently linear relationship was obtained between the ratio of variable to maximum fluorescence (F _V/F _M, 692) and the photon yield of O₂ evolution, indicating that photoinhibition affects these two variables to approximately the same extent. Treatment of leaves in a CO₂–free gas stream containing 2% O₂ and 98% N₂ under weak light (100 mol · m^–2 · s^–1) resulted in a general and fully reversible quenching of 77K fluorescence at 692 and 734 nm. In this case both F _O, 692 and F _M, 692 were invariably quenched, indicating that the quenching was caused by an increased non-radiative energy dissipation in the pigment bed. We propose that high-light treatments can have at least two different, concurrent effects on 77K fluorescence in leaves. One results from damage to the photosystem II (PSII) reaction-center complex and leads to a rise in F _O, 692; the other results from an increased non-radiative energy dissipation and leads to quenching of both F _O, 692 and F _M, 692 This general quenching had a much longer relaxation time than reported for pH-dependent quenching in algae and chloroplasts. Sun leaves, whose F _V/F _M, 692 ratios were little affected by high-light exposure in normal air, suffered pronounced photoinhibition when the exposure was made under conditions that prevent photosynthetic gas exchange (2% O₂, 0% CO₂). However, they were still less susceptible than shade leaves, indicating that the higher capacity for energy dissipation via photosynthesis is not the only cause of their lower susceptibility. The rate constant for recovery from photoinhibition was much higher in mature sun leaves than in mature shade leaves, indicating that differences in the capacity for continuous repair may in part account for the difference in their susceptibility to photoinhibition.Abbreviations and symbols kDa kilodalton - LHC-II light-harvesting chlorophyll-protein complex - PFD photon flux density (photon fluence rate) - PSI, PSII photosystem I, II - F _O, F _M, F _V instantaneous, maximum, variable fluorescence emission - absorptance - _a photon yield of O₂ evolution (absorbed light) C.I.W.-D.P.B. Publication No. 925     

Photoinhibition at chilling temperature

The effects of moderate light at chilling temperature on the photosynthesis of unhardened (acclimated to +18° C) and hardened (cold-acclimated) spinach (Spinacea oleracea L.) leaves were studied by means of fluorescence-induction measurements at 20° C and 77K and by determination of quantum yield of O₂ evolution. Exposure to 550 mol photons·m^-2·s^-1 at +4° C induced a strong photoinhibition in the unhardened leaves within a few hours. Photoinhibition manifested by a decline in quantum yield was characterized by an increase in initial fluorescence (F _o) and a decrease in variable fluorescence (F _v) and in the ratio of variable to maximum fluorescence (F _V/F _M), both at 77K and 20° C. The decline in quantum yield was more closely related to the decrease in the F _V/F _M ratio measured at 20° C, as compared with F _V/F _M at 77K. Quenching of the variable fluorescence of photosystem II was accompanied by a decline in photosystem-I fluorescence at 77K, indicating increased thermal de-excitation of pigments as the main consequence of the light treatment. All these changes detected in fluorescence parameters as well as in the quantum yield of O₂ evolution were fully reversible within 1–3 h at a higher temperature in low light. The fast recovery led us to the view that this photoinhibition represents a regulatory mechanism protecting the photosynthetic apparatus from the adverse effects of excess light by increasing thermal energy dissipation. Long-term cold acclimation probably enforces other protective mechanisms, as the hardened leaves were insensitive to the same light treatment that induced strong inhibition of photosynthesis in unhardened leaves.Abbreviations F ₀ initial fluorescence - F _M maximum fluorescence - F _V variable fluorescence (F _M-F ₀ - PFD photon flux density - PS photosystem     

On the mechanism of cytochrome oxidation in bacterial photosynthesis Quantum tunnelling effects revisited

We propose that the unique temperature dependence of the Chance-De Vault cytochrome oxidation reaction in Chromatium is not due to a transition from low-temperature nuclear tunnelling to a high-temperature activated electron transfer (ET), but rather originates from two parallel ET processes from two distinct low-potential cytochromes to the bacteriochlorophyll dimer cation. These involve a slow activationless process, which dominates at low temperatures (T 120 K) and an activated process, which is practically exclusive at high temperatures. This conjecture provides plausible nuclear and electronic coupling terms and structural data for the two cytochrome oxidation reactions.     

Quantum yields for uptake of carbon dioxide in C3 vascular plants of contrasting habitats and taxonomic groupings

The maximum quantum yields (_a,c) for CO₂ uptake in low-oxygen atmospheres were determined for 11 species of C₃ vascular plants of diverse taxa, habitat and life form using an Ulbricht-sphere leaf chamber. Comparisons were also made between tissues of varied age within species. The species examined were Psilotum nudum (L.) P. Beauv., Davallia bullata Wall. ex Hook., Cycas revoluta Thunb., Araucaria heterophylla (Salisb.) Franco, Picea abies (L.) Karst., Nerium oleander L., Ruellia humilis Nutt., Pilea microphylla (L.) Karst., Beaucarnea stricta Lem., Oplismenus hirtellus (L.) P. Beauv. and Poa annua L. Quantum yields were calculated from the initial slopes of the response of CO₂ uptake to the quantity of photons absorbed in conditions of diffuse lighting. Regression analysis of variance of the initial slopes of the response of CO₂ uptake to photon absorption failed to show any statistically significant differences between age classes within species or between the mature photosynthetic organs of different species. The constancy of _a,c was apparent despite marked variation in the light-saturated rates of CO₂ uptake within and between species. The mean _a,c was 0.093±0.003 for 11 species. By contrast, surface absorptance varied markedly between species from 0.90 to 0.60, producing proportional variation in the quantum yield calculated on an incidentlight basis. The ratio of variable to maximum fluorescence emission at 695 nm for the same tissues also failed to show any statistically significant variation between species, with a mean of 0.838±0.008. Mean values of _a,c reported here for C₃ species, in the absence of photorespiration, are higher than reported in previous surveys of vascular plants, but consistent with recent estimates of the quantum yields of O₂ evolution.Abbreviations and Symbols A rate of CO₂ uptake per unit projected area (mol · m^–2 · s^–1) - F_m the maximum fluorescence emission at 695 nm in saturating excitation light when closure of PSII reaction centres is maximal (relative units) - F_o the ground fluorescence at 695 nm when all PSII reaction centres are assumed open (relative units) - F_v the difference between F_m and F_o - J_Q rate of CO₂ uptake by the sample (nmol · s^–1) - J_Q rate of photon absorption by the sample (nmol · s^–1) - Q absorbed photon flux per unit of projected area (nmol · m^–2 · s^–1) - ₁ the light absorptance of photosynthetic organs (dimensionless) - s₁ and s'₁ the total and projected surface areas of the photosynthetic organs examined (m²) - _a,c and _i,c the quantum yields for CO₂ uptake on an absorbed- and incident-light basis, respectively (dimensionless) - _a,o the quantum yield for O₂ evolution on an absorbed-light basis (dimensionless) This work was supported by grant PI7179-BIO, FWF, Austria to H.B-N. and by a British Council travel award to S.P.L. This work was completed under the auspices of U.S. Department of Energy under Contract No. DE-AC02-76CH00016. We also thank Dr. K.J. Parkinson of PP Systems, Hitchin, UK for the loan of a prototype of a commercial integrating-sphere leaf chamber developed from our design.     

Non-invasive monitoring of photosynthetic activity and water content in forest lichens by spectral reflectance data and RGB colors from photographs

There is a need for non-invasive monitoring of temporal and spatial variation in hydration and photosynthetic activity of red-listed poikilohydric autotrophs. Here, we simultaneously recorded kinetics in RGB-colors (photos), reflectance spectra, water content, maximal (F_V/F_M), and effective quantum yield of PSII (Φ_PSII) during desiccation in foliose lichens differing in cortical characteristics and photobionts. The spectral absorbance peaks of chlorophyll a, phycocyanin, and phycoerythrin were clearly displayed at high hydration levels. Brightness and total RGB colors of the lichens strongly increased during desiccation. The normalized difference vegetation index (NDVI) efficiently estimated hydration level and Φ_PSII – a proxy for lichen photosynthesis – in all species, including threatened old forest lichens. Color and reflectance indices based on green wavelengths gave good estimates of water content in cephalo- and chlorolichens, but not in cyanolichens with a wider range of photosynthetic pigments. Due to species-specific characteristics, species-wise calibration is essential for non-invasive assessments of lichen functioning.     

Photoreactions of cytochrome b-559 and cyclic electron flow in Photosystem II of intact chloroplasts

The high potential cytochrome b-559 of intact spinach chloroplasts was photooxidized by red light with a high quantum efficiency and by far-red light with a very low quantum efficiency, when electron flow from water to Photosystem II was inhibited by a carbonyl cyanide phenylhydrazone (FCCP or CCCP). Dithiothreitol, which reacts with FCCP or CCCP, reversed the photooxidation of cytochrome b-559 and restored the capability of the chloroplasts to photoreduce CO₂ showing that the FCCP/CCCP effects were reversible. The quantum efficiency of cytochrome b-559 photooxidation by red or far-red light in the presence of FCCP was increased by 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone which blocks oxidation of reduced plastoquinone by Photosystem I. When the inhibition of water oxidation by FCCP or CCCP was decreased by increased light intensities, previously photooxidized cytochrome b-559 was reduced. Red light was much more effective in photoreducing oxidized high potential cytochrome b-559 than far-red light. The red/far-red antagonism in the redox state of cytochrome b-559 is a consequence of the different sensitivity of the cytochrome to red and far-red light and does not indicate that the cytochrome is in the main path of electrons from water to NADP. Rather, cytochrome b-559 acts as a carrier of electrons in a cyclic path around Photosystem II. The redox state of the cytochrome was shifted to the oxidized side when electron transport from water became rate-limiting, while oxidation of water and reduction of plastoquinone resulted in its shifting to the reduced side.     

Quantum yield and rate of formation of the carotenoid triplet state in photosynthetic structures

The formation of the triplet state of carotenoids (detected by an absorption peak at 515 nm) and the photo-oxidation of the primary donor of Photosystem II, P-680 (detected by an absorption increase at 820 nm) have been measured by flash absorption spectroscopy in chloroplasts in which the oxygen evolution was inhibited by treatment with Tris. The amount of each transient form has been followed versus excitation flash intensity (at 590 or 694 nm). At low excitation energy the quantum yield of triplet formation (with the Photosystem II reaction center in the state Q⁻) is about 30% that of P-680 photo-oxidation. The yield of carotenoid triplet formation is higher in the state Q⁻ than in the state Q, in nearly the same proportion as chlorophyll a fluorescence. It is concluded that, for excited chlorophyll a, the relative rates of intersystem crossing to the triplet state and of fluorescence emission are the same in vivo as in organic solvent. At high flash intensity the signal of P-680⁺ completely saturates, whereas that of carotenoid triplet continues to increase.

The rate of triplet-triplet energy transfer from chlorophyll a to carotenoids has been derived from the rise time of the absorption change at 515 nm, in chloroplasts and in several light-harvesting pigment-protein complexes. In all cases the rate is very high, around 8 · 10⁷ s⁻¹ at 294 K. It is about 2–3 times slower at 5 K. The transitory formation of chlorophyll triplet has been verified in two pigment-protein complexes, at 5 K.