Primary Electron Donor Of Photosystem Research Articles

Photo-Oxidation of P740, the Primary Electron Donor in Photosystem I from Acaryochloris marina

Fourier transform infrared spectroscopy (FTIR) difference spectroscopy in combination with deuterium exchange experiments has been used to study the photo-oxidation of P740, the primary electron donor in photosystem I from Acaryochloris marina. Comparison of (P740 +-P740) and (P700 +-P700) FTIR difference spectra show that P700 and P740 share many structural similarities. However, there are several distinct differences also: 1), The (P740 +-P740) FTIR difference spectrum is significantly altered upon proton exchange, considerably more so than the (P700 +-P700) FTIR difference spectrum. The P740 binding pocket is therefore more accessible than the P700 binding pocket. 2), Broad, “dimer” absorption bands are observed for both P700 + and P740 +. These bands differ significantly in substructure, however, suggesting differences in the electronic organization of P700 + and P740 +. 3), Bands are observed at 2727(−) and 2715(−) cm −1 in the (P740 +-P740) FTIR difference spectrum, but are absent in the (P700 +-P700) FTIR difference spectrum. These bands are due to formyl CH modes of chlorophyll d. Therefore, P740 consists of two chlorophyll d molecules. Deuterium-induced modification of the (P740 +-P740) FTIR difference spectrum indicates that only the highest frequency 13 3 ester carbonyl mode of P740 downshifts, indicating that this ester mode is weakly H-bonded. In contrast, the highest frequency ester carbonyl mode of P700 is free from H-bonding. Deuterium-induced changes in (P740 +-P740) FTIR difference spectrum could also indicate that one of the chlorophyll d 3 1 carbonyls of P740 is hydrogen bonded.

Mutation induced modulation of hydrogen bonding to P700 studied using FTIR difference spectroscopy.

Site-directed mutagenesis in combination with Fourier transform infrared difference spectroscopy has been used to study how hydrogen bonding modulates the electronic and physical organization of P700, the primary electron donor in photosystem I. Wild-type PS I particles from Chlamydomonas reinhardtii and a mutant in which ThrA739 is changed to alanine [TA(A739) mutant] were studied. ThrA739 is thought to provide a hydrogen bond to the chlorophyll-a' molecule of P700 (the two chlorophylls of P700 (P700(+)) will be called P(A) and P(B) (P(A)(+) and P(B)(+))). The mutation considerably alters the (P700(+)-P700) FTIR difference spectra. However, we were able to describe all of the mutation induced changes in the difference spectra in terms of difference band assignments that were proposed recently (Hastings, G., Ramesh, V. M., Wang, R., Sivakumar, V. and Webber, A. (2001) Biochemistry 40, 12943-12949). Upon comparison of mutant and wild type (P700(+)-P700) FTIR difference spectra, it is shown that (1) the 13(3) ester carbonyl modes of P(A) and P(B) are unaltered upon mutation of ThrA739 to alanine. (2) The 13(3) ester carbonyl modes of P(A)(+)/P(B)(+) upshift/downshift upon mutation. These oppositely directed shifts indicate that the mutation modifies the charge distribution over the pigments in the P700(+) state, with charge on P(B) being relocated onto P(A). We also show that the 13(1) keto carbonyl mode of P(B)/P(B)(+) is unaltered/downshifted upon mutation, as is expected for the above-described mutation induced charge redistribution in P700(+). Although the 13(3) ester carbonyl modes of the chlorophylls of P700 in the ground state are unaltered upon mutation, the 13(1) keto carbonyl mode of P(A) upshifts upon mutation, as does the 13(1) keto carbonyl mode of P(A)(+). For P700 in the ground state, bands that we associate with HisA676/HisB656 upshift/downshift upon mutation. For the P700(+) state, bands that we associate with HisA676/HisB656 also upshift/downshift upon mutation. These observations are also consistent with the notion that the mutation leads to the charge on P(B)(+) being relocated onto P(A)(+). In addition, we suggest that a hydrogen bond to the 13(1) keto carbonyl of P(A) is still present in the TA(A739) mutant, probably mediated through an introduced water molecule.

The two histidine axial ligands of the primary electron donor chlorophylls (P700) in photosystem I are similarly perturbed upon P700+ formation.

The extent of delocalization of the positive charge in the oxidized dimer of chlorophyll (Chl) constituting P700, the primary electron donor of photosystem I (PSI), has been investigated by analyzing the perturbation upon P700(+) formation of infrared (IR) vibrational modes of the two His axial ligands of the two P700 Chl molecules. Fourier transform IR (FTIR) difference spectra of the photooxidation of P700 in PSI core complexes isolated from Synechocystis sp. PCC 6803 isotopically labeled either globally with (15)N or more specifically with (13)C on all the His residues reveal isotopic shifts of a differential signal at 1102/1108 cm(-)(1). This signal is assigned to a downshift upon P700(+) formation of the predominantly C(5)-Ntau imidazole stretching mode of His residue(s). The amplitude of this signal is reduced by approximately half in FTIR spectra of Synechocystis mutants in which His PsaB 651, the axial ligand to one of the two Chl molecules in P700, is replaced by Cys, Gln, or Leu. These observations provide further evidence that the positive charge in P700(+) is essentially delocalized over the two Chl molecules, in agreement with a previous FTIR study in which the frequency of the vibrational modes of the 9-keto and 10a-ester C=O groups of the two Chl's in P700, P700(+), and (3)P700 were firmly established for the first time [Breton, J., et al. (1999) Biochemistry 38, 11585-11592]. Only limited perturbations of the amplitude and frequency of the 9-keto and 10a-ester C=O bands of the P700 Chl are elicited by the mutations. On the basis of comparable mutational studies of the primary electron donor in purple bacteria, these perturbations are attributed to small molecular rearrangements of the Chl macrocycle and substituents caused by the repositioning of the P700 dimer in the new protein cavity generated by the mutations. It is proposed that the perturbation of the FTIR spectra upon mutation of a His axial ligand of the P700 Chl recently reported in Chlamydomonas reinhardtii [Hastings, G., et al. (2001) Biochemistry 40, 12943-12949] can be explained by the same effect without the need for a new assignment of the C=O bands of P700. The distribution of charge/spin in P700(+) and (3)P700 determined by FTIR spectroscopy is discussed in relation with the contrasting interpretations derived from recent magnetic resonance experiments.

P680, the primary electron donor of photosystem II

The primary electron donor of photosystem II is a special form of chlorophyll a known as P680. Its detection and subsequent biophysical characterisation has relied heavily on the technique of flash photolysis of Norrish and Porter [Nature 164 (1949) 658] and on the physical principles which emerged from photochemical studies of isolated chlorophyll a using this technique. When oxidised the P680 radical has a midpoint redox potential estimated to be 1.17 V or more which is needed to drive the oxidising reactions of the water-splitting process. Such a high oxidising potential dictates special properties of P680 which are discussed in terms of robustness and structural organisation of photosystem II. Of particular importance has been the recent finding that P680 is not a ‘special pair’ of chlorophyll molecules as is the case for the primary electron donors of other types of photosynthetic reaction centres. Instead P680 is composed of a cluster of four weakly coupled monomeric chlorophylls which together with the local protein environment enables this primary donor to generate a redox potential capable of oxidising water.

Time-Resolved Step-Scan Fourier Transform Infrared and Visible Absorption Difference Spectroscopy for the Study of Photosystem I

Step-scan Fourier transform infrared and visible absorption difference spectroscopy, with nanosecond to millisecond time resolution, has been applied to the study of photosystem I particles from photosynthetic oxygen-evolving organisms. In particular, time-resolved infrared (1800–1200 cm−1) and visible (680–850 nm) difference spectra associated with flash induced oxidation of the primary electron donor in photosystem I particles from Synechocystis sp. 6803 and Acarychloris marina are presented. These spectra are compared directly to static, photoaccumulated Fourier transform infrared and visible difference spectra from both species. The detailed bonding interactions of P740, the primary donor in photosystem I particles from Acarychloris marina, are very similar to that found for P700, the primary donor in photosystem I particles from Synechocystis sp. 6803. P740 consists of at least two chlorophyll molecules, both of which are probably chlorophyll-d. Combined time-resolved step-scan Fourier transform infrared and visible absorption difference spectroscopy is shown to be a powerful tool for the study of single molecular bond dynamics in proteins as large as 0.4 mega-Daltons.

Secondary stabilization reactions and proton-coupled electron transport in photosystem II investigated by electroluminescence and fluorescence spectroscopy.

The oxidized primary electron donor in photosystem II, P(680)(+), is reduced in several phases, extending over 4 orders of magnitude in time. Especially the slower phases may reflect the back-pressure exerted by water oxidation and provide information on the reactions involved. The kinetics of secondary electron-transfer reactions in the microseconds time range after charge separation were investigated in oxygen-evolving thylakoids suspended in H2O or D2O. Flash-induced changes of chlorophyll fluorescence yield and electric field-induced recombination luminescence were decomposed into contributions from oxidation states S(0), S(1), S(2), and S(3) of the oxygen-evolving complex and interpreted in terms of stabilization kinetics of the initial charge-separated state S(j)Y(Z)P(680)(+)Q(A)(-)Q(B). In approximately 10% of the centers, only charge recombination took place. Otherwise, no static heterogeneity was involved in the microsecond reduction of P(680)(+) by Y(Z) (stabilization) or Q(A)(-) (recombination). The recombination component in active centers occurs mainly upon charge separation in S(3), and, in the presence of D2O, in S(2) as well and is tentatively attributed to the presence of Y(Z)(ox)S(j-1) in equilibrium with Y(Z)S(j). A 20-30 micros stabilization occurs in all S-states, but to different extents. Possible mechanisms for this component are discussed. D2O was found to decrease: (i) the rate of the reaction Y(Z)(ox)S(1) to Y(Z)S(2), (ii) the equilibrium constant between P680(+)Y(Z)S(2) and P(680)Y(Z)(ox)S(2), (iii) the rate of the slow phase of P(680)(+) reduction for the S(3) --> S(0) transition, and (iv) the rate of electron transfer from Q(A)(-) to Q(B) /Q(B)(-). The increased 'miss probability' in D2O is due to (iii).

Normal-phase HPLC separation of possible biosynthetic intermediates of pheophytin a and chlorophyll a'.

Normal-phase HPLC conditions have been developed for separating the C17(3) isoprenoid isomers, which are expected to be formed as biosynthetic intermediates of chlorophyll (Chl) a, Chl a' (C13(2)-epimer of Chl a), pheophytin (Pheo) a and protochlorophyll (PChl). The application of these conditions to pigment composition analysis of greening etiolated barley leaves allowed us to detect, for the first time, the C17(3) isomers of Chl a', a possible constituent of the primary electron donor of photosystem (PS) I, P700, and those of Pheo a, the primary electron acceptor of PS II, in the very early stage of greening. The C17(3) isomer distribution patterns were approximately the same between Chl a and Chl a', but significantly different between Pheo a and Chl a', probably reflecting the similarity and difference, respectively, in the biosynthetic pathways of these pigment pairs.

FTIR study of the primary electron donor of photosystem I (P700) revealing delocalization of the charge in P700(+) and localization of the triplet character in (3)P700.

The effect of global (15)N or (2)H labeling on the light-induced P700(+)/P700 FTIR difference spectra has been investigated in photosystem I samples from Synechocystis at 90 K. The small isotope-induced frequency shifts of the carbonyl modes observed in the P700(+)/P700 spectra are compared to those of isolated chlorophyll a. This comparison shows that bands at 1749 and 1733 cm(-)(1) and at 1697 and 1637 cm(-)(1), which upshift upon formation of P700(+), are candidates for the 10a-ester and 9-keto C=O groups of P700, respectively. A broad and relatively weak band peaking at 3300 cm(-)(1), which does not shift upon global labeling or (1)H-(2)H exchange, is ascribed to an electronic transition of P700(+), indicating that at least two chlorophyll a molecules (denoted P(1) and P(2)) participate in P700(+). Comparisons of the (3)P700/P700 FTIR difference spectrum at 90 K with spectra of triplet formation in isolated chlorophyll a or in RCs from photosystem II or purple bacteria identify the bands at 1733 and 1637 cm(-)(1), which downshift upon formation of (3)P700, as the 10a-ester and 9-keto C=O modes, respectively, of the half of P700 that bears the triplet (P(1)). Thus, while the P(2) carbonyls are free from interaction, both the 10a-ester and the 9-keto C=O of P(1) are hydrogen bonded and the latter group is drastically perturbed compared to chlorophyll a in solution. The Mg atoms of P(1) and P(2) appear to be five-coordinated. No localization of the triplet on the P(2) half of P700 is observed in the temperature range of 90-200 K. Upon P700 photooxidation, the 9-keto C=O bands of P(1) and P(2) upshift by almost the same amount, giving rise to the 1656(+)/1637(-) and 1717(+)/1697(-) cm(-)(1) differential signals, respectively. The relative amplitudes of these differential signals, as well as of those of the 10a-ester C=O modes, appear to be slightly dependent on sample orientation and temperature and on the organism used to generate the P700(+)/P700 spectrum. If it is assumed that the charge density on ring V of chlorophyll a, as measured by the perturbation of the 10a-ester or 9-keto C=O IR vibrations, mainly reflects the spin density on the two halves of the oxidized P700 special pair, a charge distribution ranging from 1:1 to 2:1 (in favor of P(2)) is deduced from the measurements presented here. The extreme downshift of the 9-keto C=O group of P(1), indicative of an unusually strong hydrogen bond, is discussed in relation with the models previously proposed for the PSI special pair.

Modulation of quantum yield of primary radical pair formation in photosystem II by site-directed mutagenesis affecting radical cations and anions.

Pigment-protein interactions play a significant role in determining the properties of photosynthetic complexes. Site-directed mutants of Synechocystis PCC 6803 have been prepared which modify the redox potential of the primary radical pair anion and cation. In one set of mutants, the environment of P680, the primary electron donor of Photosystem II, has been modified by altering the residue at D1-His198. It has been proposed that this residue is an axial ligand to the magnesium cation. In the other set, the D1-Gln130 residue, which is thought to interact with the C9-keto group of the pheophytin electron acceptor, has been changed. The effect of these mutations is to alter the free energy of the primary radical pair state, which causes a change in the equilibrium between excited singlet states and radical pair states. We show that the free energy of the primary radical pair can be increased or decreased by modifications at either the D1-His198 or the D1-Gln130 sites. This is demonstrated by using three independent measures of quantum yield and equilibrium constant, which exhibit a quantitative correlation. These data also indicate the presence of a fast nonradiative decay pathway that competes with primary charge separation. These results emphasize the sensitivity of the primary processes of PS II to small changes in the free energy of the primary radical pair.

Submillimeter High-Field EPR Studies of the Primary Donor in Plant Photosystem I P700•+

In this present study we report millimeter and submillimeter high-field CW EPR spectra of P700•+, the primary electron donor in photosystem I. The data shown are the first well-resolved chlorophyll radical spectra observed without prior deuteration. The signal was generated from isolated plant photosystem I (in both digitonin and Triton-X100 preparations) by photooxidation. At 325 GHz and higher frequencies, the spectrum was resolvable into the three principal components of its g-matrix. They were obtained to high accuracy by spectral simulation and were found to be gxx = 2.003 17(±7 × 10-5), gyy = 2.002 64(±7 × 10-5), gzz = 2.002 26(±7 × 10-5) at 40 K and gxx = 2.003 07(±7 × 10-5), gyy = 2.002 60(±7 × 10-5), gzz = 2.002 26(±7 × 10-5) at 200 K. These values indicate a temperature dependence of the g-values in P700•+ that was measured over a temperature range of 5−260 K using both preparations. The results showed that gxx changed with temperature while gyy and gzz remain constant within the error margin, s...

Stabilization of oxygen evolution and primary electron transport reactions in photosystem II against heat stress with glycinebetaine and sucrose

The protective action of co-solutes, such as sucrose and glycinebetaine, againts the thermal inactivation of photosystem II function was studied in untreated and Mn-depleted photosystem II preparations. It was shown that, in addition to the reactions that depend on the oxygen-evolving activity of the photosystem, those that implicate more intimately the reaction center itself are protected by high concentrations of osmolytes. However, the temperature required to inhibit oxygen evolution totally in the presence of osmolytes is lower than that required to eliminate reactions, such as P680 (primary electron donor in photosystem II) photo-oxidation and pheophytin photoreduction, which only involve charge separation and primary electron transport processes. The energy storage measured from the thermal dissipation yield during photoacoustic experiments and the yield of variable fluorescence are also protected to a significant degree (up to 30%) at temperatures at which oxygen evolution is totally inhibited. It is suggested that a cyclic electron transport reaction around photosystem II may be preserved under these conditions and may be responsible for the energy storage measured at relatively high temperatures. This interpretation is also supported by thermoluminescence data involving the recombination between reduced electron acceptors and oxidized electron donors at −30 and −55 °C. The data also imply that a high concentration of osmolyte allows the stabilization of the photosystem core complex together with the oxygen-evolving complex. The stabilization effect is understood in terms of the minimization of protein-water interactions as proposed by the theory of Arakawa and Timasheff ( Biophys. J., 47 (1985) 411–414).

Characterization of the primary electron donor of photosystem I, P700, by electrochemistry and Fourier transform infrared (FTIR) difference spectroscopy

Abstract The redox reactions of P700, the primary electron donor of photosystem I (PS I), have been analyzed by electrochemistry and Fourier transform infrared (FTIR) spectroscopy in two different preparations from the cyanobacterium Synechocystis PCC 6803 yielding monomeric and trimeric PS I complexes, respectively. Reversible and quantitative oxidation and rereduction of P700 has been achieved at surface-modified electrodes in an optically and infrared transparent thin-layer electrochemical (OTTLE) cell. The midpoint potential ( E m = +260 ± 10 mV vs. Ag/AgCl/3 M KCl) determined from these titrations, either by monitoring the electrochemically-induced oxidation/reduction of the primary electron donor P700 at 700 nm or by monitoring the amplitude of the flash-induced absorbance change at 707 nm as a function of the applied potential, agree between monomeric and trimeric PS I, demonstrating that protein-protein contact has very little impact on the redox properties of P700. Using electrochemical oxidation and rereduction, the vibrational IR difference spectra P700 ·+ − P700 could be generated. In the 1800−1200 cm −1 spectral region, highly structured IR difference spectra were obtained. The IR difference bands titrate in unison and are fully reversible upon application of a reducing potential. The midpoint potential measured by titration of the absorbance difference of two prominent bands, at 1714 and 1696 cm −1 ( E m = +260 ± 10 mV) corresponds to the values determined by titration of the electronic transition of P700. In contrast to previously obtained FTIR difference spectra for light-induced charge separation (E. Nabedryk, M. Leonhard, W. Mantele and J. Breton, Biochemistry, 29 (1990) 3242; G. MacDonald, K.A. Bixby and B.A. Barry, Proc. Natl. Acad. Sci. USA, 90 (1993) 11024) which include contributions from the primary electron donor and the acceptor side, the electrochemical generated FTIR difference spectra exclusively reflect the molecular changes at P700 and its protein site upon removal of an electron. Nevertheless, they largely agree with the difference spectra of the charge-separated state. In the 1800-1680 cm −1 range, the IR difference signals are attributed to the keto and ester C=O modes of P700. Between 1700 and 1680 cm −1 , shifts of bands and changes of band intensities for photosystem I particles suspended in 2 H 2 O reveal contributions of amide I modes. Further effects of 2 H 2 O/ 1 H 2 O exchange are observed between 1600 and 1500 cm −1 , and around 1400 cm −1 . Between 1750 cm −1 and approximately 3000 cm −1 , no FTIR difference signals were detected. In particular, no broad but strong absorbance changes were detected around 2600 cm −1 , where a low-energy electronic transition, characteristic for the dimeric nature of P and P + , was detected for P865 in bacterial reaction centers (J. Breton, E. Nabedryk and W. Parson, Biochemistry, 31 (1992) 7503). This indicates that the positive charge on P700 ·+ is rather localized on one of the chlorophylls.

Identification of histidine as an axial ligand to P700+.

The primary electron donor in photosystem I (PSI), P700, is thought to be a dimeric Chl a species. Neither the electronic nor geometric structure of the cation radical is clearly understood. Magnetic resonance studies have indicated that the unpaired electron in P700+ is delocalized asymmetrically over the Chl dimer; however, the axial ligand to the central Mg2+ is not known. The recent development of a histidine tolerant mutant of Synechocystis PCC 6803 has allowed us to use a combination of isotopic labeling and electron nuclear double resonance (ENDOR) spectroscopy to show the first definitive spectroscopic evidence of a histidine ligand to P700+. Peaks split symmetrically about the 15N Larmor frequency corresponding to an isotropic hyperfine coupling of 0.64 MHz were observed in the ENDOR spectra from P700+ globally labeled with 15N and specifically labeled with [15N]histidine. These peaks disappeared in "reverse" labeled samples in which all nitrogens are 15N except those of histidine, which contains natural abundance 14N. The dipolar contribution to the hyperfine coupling was determined by using electron spin echo envelope modulation spectroscopy (ESEEM). Numerical simulations of the ESEEM data suggest that the coupling is primarily isotropic and that the histidine is directly coordinated to the central Mg2+ of P700+. Taken together, these data are supportive of a model of P700+ in which the excited state molecular orbital makes a significant contribution to the electronic structure of the radical. Moreover, the methodology developed in this work can be extended to examine the magnetic properties of axial ligands in a variety of biologically relevant porphyrin/chlorin systems.

Site-directed mutations affecting the spectroscopic characteristics and midpoint potential of the primary donor in photosystem I.

Photosystem I is a member of the iron-sulfur center or type I reaction centers. The primary electron donor in photosystem I is a chlorophyll a dimer termed P700. The biophysical properties of P700 are well understood, but the protein environment that gives it such unique properties is unknown. We have characterized site-directed mutants of the photosystem I reaction center protein PsaB and identified an amino acid, His-656, that interacts closely with one of the P700 chlorophylls. Mutation of His-656 to Asn or Ser increases the oxidation midpoint potential of P700/P700+. by 40 mV. The P700/P700+. optical difference spectra show the appearance of a new bleaching band at 667 nm. Electron nuclear double resonance spectroscopy indicates a significant increase in the hyperfine coupling corresponding to methyl protons at position 12 of the spin carrying chlorophyll a of P700+. The implication of these results to current structural models of the photosystem I reaction center is discussed.

Purification and spectroscopic characterization of photosystem II reaction center complexes isolated with or without Triton X-100.

The pigment composition of the isolated photosystem II reaction center complex in its most stable and pure form currently is a matter of considerable debate. In this contribution, we present a new method based on a combination of gel filtration chromatography and diode array detection to analyze the composition of photosystem II reaction center preparations. We show that the method is very sensitive for the detection of contaminants such as the core antenna protein CP47, pigment-free and denatured reaction center proteins, and unbound chlorophyll and pheophytin molecules. We also present a method by which the photosystem II reaction center complex is highly purified without using Triton X-100, and we show that in this preparation the contamination with CP47 is less than 0.1%. The results strongly indicate that the photosystem II reaction center complex in its most stable and pure form binds six chlorophyll a, two pheophytin a, and two beta-carotene molecules and that the main effect of Triton X-100 is the extraction of beta-carotene from the complex. Analysis of 4 K absorption and emission spectra indicates that the spectroscopic properties of this preparation are similar to those obtained by a short Triton X-100 treatment. In contrast, preparations obtained by long Triton X-100 treatment show decreased absorption of the shoulder at 684 nm in the 4 K absorption spectrum and an increased number of pigments that trap excitation energy at very low temperatures. We conclude that the 684 nm shoulder in the 4 K absorption spectrum should at least in part be attributed to the primary electron donor of photosystem II.

Subpicosecond Transient Absorption Difference Spectroscopy on the Reaction Center of Photosystem II: Radical Pair Formation at 77 K

To further characterize the processes of charge separation and excitation energy transfer in the isolated photosystem II reaction center complex (the D1-D2-cytochrome b-559 complex), subpicosecond absorption difference spectra were measured at 77 K. The preparations were excited with a repetition rate of 30 Hz by laser pulses with a duration of 300-400 fs at 688, 681, 672, and 670.5 nm. Upon excitation at 688 and 681 nm, a biphasic decay of the induced signals with time constants of 1-2 ps and 80-100 ps was observed. The 1-2 ps component is interpreted as the intrinsic charge separation time from the singlet-excited state of P680, the primary electron donor of photosystem II. The 80-100 ps time is interpreted as charge separation limited by slow energy transfer from a trap state in the photosystem II reaction center degenerate with P680, earlier identified by low-temperature fluorescence experiments. In addition, upon excitation at 672 and 670.5 nm energetically downhill energy transfer processes of 400-500 fs and nearly 14 ps were observed, which according to the associated spectral changes, originate from states in the reaction center absorbing at different energies. 40 refs., 6 figs.

Copper and photosystem II: A controversial relationship

Copper is an essential micronutrient for higher plants and algae and has a direct impact on photosynthesis. It is a constituent of the primary electron donor in photosystem I, the Cu‐protein plastocyanin. Many authors have also described Cu as a constituent of photosystem II (PSII). However, high Cu concentrations inhibit the photosynthetic electron transport, especially in PSII. In addition, both Cu deficiency and Cu toxicity interfere with pigment and lipid biosynthesis and, consequently, with chloroplast ultrastructure thus negatively influencing the photosynthetic efficiency.In this review, the different functions proposed for the metal in PSII are reviewed. With reference to the effect of toxic Cu on PSII, the polemic results concerning its mechanism of action and Cu‐binding sites are discussed. Other effects of Cu toxicity and Cu deprivation on the thylakoid membrane are also briefly described.

Does slow energy transfer limit the observed time constant for radical pair formation in photosystem II reaction centers?

We have used spectrally photoselective femtosecond transient absorption spectroscopy on photosystem II reaction centers to show that there are at least two pools of chlorin molecules/states which can transfer excitation energy to P680, the primary electron donor in photosystem II. It has previously been shown that one chlorin pool equilibrates with P680 in 100 fs [Durrant et al. (1992) Proc. Natl. Acad. Sci. U.S.A. 89, 11632-11636], and we report here the observation of energy transfer from a second more weakly coupled chlorin pool. The effect of the weakly coupled pool is to increase the apparent time constant for radical pair formation from 21 ps when P680 is selectively excited to 27 ps when the accessory chlorins are excited. We conclude that it is possible to observe both radical pair formation somewhat slowed by an energy transfer step and radical pair formation not limited by this slow energy transfer, depending upon which chromophores are initially excited. These observations provide evidence that when using photoselective excitation of P680, the observed 21 ps time constant for radical pair formation is not limited by a slow energy transfer step.

Light-induced Fourier transform infrared spectrum of the cation radical P680+

The structure of the primary electron donor of photosystem II, P680, is still under debate. It is not decided if it is composed of a chlorophyll (Chl) monomer or dimer. In this study, Fourier transform infrared (FTIR) spectroscopy was used to analyze the changes in the vibration modes occurring upon photooxidation of P680 in a Mn-depleted PS II preparation. It is demonstrated that illumination of the above in the presence of artificial electron acceptors results in a light-minus-dark absorbance change typical of the formation ofP680+. The light-minus-dark difference FTIR spectrum obtained under similar conditions is characterized by two negative peaks located at 1694 and 1652 or 1626 cm−1 that can be assigned to the 9-keto groups of the P680 Chl, the latter band being indicative of a strongly associated group. These vibrations are shifted to 1714 and 1676 cm−1, respectively, in the positive features of the difference spectrum attributed to P680+. The occurrence of two pairs of bands attributed to 9-keto groups is discussed in terms of P680 being formed of a Chl dimer.

Transient electron paramagnetic resonance of the triplet state of P700 in photosystem I: Evidence for triplet delocalization at room temperature

Spin-polarized EPR spectra of the triplet state of P700, the primary electron donor in photosystem I (PS I), have been measured for the first time at room temperature. The measurements were performed on intact PS I from Synechococcus sp. after prereduction of all iron-sulfur centers and on vitamin K1 depleted PS I from Synechocystis 6803. The two preparations give similar spectra with a polarization pattern which indicates that the triplet state is formed via recombination of a radical pair. The axial and nonaxial zero-field splitting (zfs) parameters are found to be magnitude of D = (284 +/- 15) x 10(-4) cm-1 and magnitude of E = (22 +/- 3) x 10(-4) cm-1, respectively. The E-value is 42% smaller than in monomeric chlorophyll a, while the D-value is nearly the same. Measurements of the Synechocystis 6803 sample at 4.5 K yielded zfs parameters which are identical with those of the chlorophyll monomer, in agreement with previous results. In order to explain this behavior, it is proposed that the triplet excitation is delocalized over the two halves of a chlorophyll dimer at room temperature but appears localized on one half at low temperature. The observed zfs parameters are obtained if (1) the magnetic z-axes of the two chlorophylls are collinear, (2) the magnetic y-axes (and x-axes) of the two chlorophylls make an angle of approximately 55 degrees with each other, and (3) the admixture of charge-transfer states to 3P700 is negligible.(ABSTRACT TRUNCATED AT 250 WORDS)