Excitonic interactions in the reaction centre of photosystem II studied by using circular dichroism

Changes in excitonic interactions of photosystem II (PSII) reaction centre (RC) pigments upon light-induced oxidation of primary donor (P680) or reduction of primary acceptor (pheophytin (Pheo)) were analysed using circular dichroism (CD). The CD spectrum of PSII RC shows positive bands at 417, 435 and 681 and negative bands at 447 and 664 nm. Oxidation of the primary donor by illuminating the sample in the presence of silicomolybdate resulted in nearly symmetric decrease of CD amplitudes at 664 and 684 nm. In the Soret region, the maximum bleaching of CD signal was detected at 449 and 440 nm. Accumulation of reduced Pheo in the presence of dithionite brought about much lower changes in CD amplitudes than P680 oxidation. In this case, only a small asymmetric bleaching at 680 and 668 nm in the red region and a bleaching at 445, 435 and 416 nm in the Soret region has been detected. Therefore, we suppose that the contribution of the Pheo of the primary acceptor to the total CD signal of RC is negligible. In contrast to the oxidation of primary donor, the light-induced change in the CD spectrum upon primary acceptor reduction was strongly temperature-dependent. The reversible CD bleaching was completely inhibited below 200 K, although the reduced Pheo was accumulated even at a temperature of 77 K. Since the temperature does not influence the excitonic interaction, the temperature dependence of the CD changes upon Pheo reduction does not support the model of Pheo excitonically interacting with the other chlorophylls (Chl) of the RC. We propose that Pheo should not be considered as a part of a multimer model.     

Primary light-energy conversion in tetrameric chlorophyll structure of photosystem II and bacterial reaction centers: II. Femto- and picosecond charge separation in PSII D1/D2/Cyt b559 complex

In Part I of the article, a review of recent data on electron-transfer reactions in photosystem II (PSII) and bacterial reaction center (RC) has been presented. In Part II, transient absorption difference spectroscopy with 20-fs resolution was applied to study the primary charge separation in PSII RC (DI/DII/Cyt b 559 complex) excited at 700 nm at 278 K. It was shown that the initial electron-transfer reaction occurs within 0.9 ps with the formation of the charge-separated state P680(+)Chl(D1)(-), which relaxed within 14 ps as indicated by reversible bleaching of 670-nm band that was tentatively assigned to the Chl(D1) absorption. The subsequent electron transfer from Chl(D1)(-) within 14 ps was accompanied by a development of the radical anion band of Pheo(D1) at 445 nm, attributable to the formation of the secondary radical pair P680(+)Pheo(D1)(-). The key point of this model is that the most blue Q(y) transition of Chl(D1) in RC is allowing an effective stabilization of separated charges. Although an alternative mechanism of charge separation with Chl(D1)* as a primary electron donor and Pheo(D1) as a primary acceptor can not be ruled out, it is less consistent with the kinetics and spectra of absorbance changes induced in the PSII RC preparation by femtosecond excitation at 700 nm.     

Photodamage to Pigment in the Photosystem Ⅱ Reaction Center D1/D2/Cytochrome b559 Complex

Strong light (800 μmol photons/m2 per s)-induced bleaching of the pigment in the isolated photosystem Ⅱ reaction center (PSII RC) under aerobic conditions (in the absence of electron donors or acceptors) was studied using high-pressure liquid chromatography (HPLC), absorption spectra, 77K fluorescence spectra and resonance Raman spectra. Changes in pigment composition of the PSll RC as determined by HPLC after light treatment were as follows: with increasing illumination time chlorophyll (Chi) a and β-carotene (β-car)content decreased. However, decreases in pheophytin (Pheo) could not be observed because of the mixture of the Pheo formed by degraded chlorophyll possibly. On the basis of absorption spectra, it was determined that, with a short time of illumination, the initial bleaching occurred maximally at 680 nm but that with increasing illumination time there was a blue shift to 678 nm. It was suggested that P680 was destroyed initially, followed by the accessory chlorophyll. The activity of P680 was almost lost after 10 min light treatment. Moreover, the bleaching of Pheo and β-car was observed at the beginning of illumination.After illumination, the fluorescence emission intensity changed and the fluorescence maximum blue shifted,showing that energy transfer was disturbed. Resonance Raman spectra of the PSII RC excited at 488.0 and 514.5 nm showed four main bands, peaking at 1 527 cm-1 (υ1), 1 159 cm-1 (υ2), 1 006 cm-1 (υ3), 966 cm-1 (υ4) for 488.0 nm excitation and 1 525 cm-1 (υ1), 1 159 cm-1 (υ2), 1 007 cm-1 (υ3), 968 cm-1 (υ4) for 514.5 nm excitation.It was confirmed that two spectroscopically different β-car molecules exist in the PSII RC. After light treatment for 20 min, band positions and bandwidths were unchanged. This indicates that carotenoid configuration is not the parameter that regulates photoprotection in the PSII RC.     

Primary Events, Energy Transfer, and Reactions in Photosynthetic Units: The Relationship between P-680 and C-550

The published reports of flash-induced absorbance changes in the 680-690 nm spectral region, which have been attributed to bleaching of the primary reaction center chlorophyll of photosystem II (PSII) P-680, are discussed in light of what is known about the primary electron acceptor of PSII, C-550. The question of whether the fluorescence yield changes, which accompany the photoreduction of C-550, might influence the measurements of chlorophyll bleaching is examined. The responses attributed to P-680 and their relationship to C-550 indicate that, if the absorbance measurements are valid, P-680 probably functions as the primary electron donor to PSII rather than as a photochemical sensitizer of the primary redox reaction.     

Primary radical ion pairs in photosystem II core complexes

Ultrafast absorption spectroscopy with 20-fs resolution was applied to study primary charge separation in spinach photosystem II (PSII) reaction center (RC) and PSII core complex (RC complex with integral antenna) upon excitation at maximum wavelength 700–710 nm at 278 K. It was found that the initial charge separation between P680* and Chl_D1 (Chl-670) takes place with a time constant of ～1 ps with the formation of the primary charge-separated state P680* with an admixture of: P680^*(1?δ) (P680^δ+1Chl _D1 ^δ? ), where δ ～ 0.5. The subsequent electron transfer from P680^δ+Chl _D1 ^δ? to pheophytin (Pheo) occurs within 13 ps and is accompanied by a relaxation of the absorption band at 670 nm (Chl _D1 ^δ? ) and bleaching of the Pheo_D1 bands at 420, 545, and 680 nm with development of the Pheoband at 460 nm. Further electron transfer to Q_A occurs within 250 ps in accordance with earlier data. The spectra of P680⁺ and Pheo^? formation include a bleaching band at 670 nm; this indicates that Chl-670 is an intermediate between P680 and Pheo. Stimulated emission kinetics at 685 nm demonstrate the existence of two decaying components with time constants of ～1 and ～13 ps due to the formation of P680^δ+Chl _D1 ^δ? and P680⁺Pheo _D1 ^? , respectively.     

Photodamage to Pigment in the Photosystem Ⅱ Reaction Center D1/D2/Cytochrome b559 Complex

Strong light （800μmol photons/m^2 per s）-induced bleaching of the pigment in the isolated photosystem Ⅱ reaction center （PSII RC） under aerobic conditions （in the absence of electron donors or acceptors） was studied using high-pressure liquid chromatography （HPLC）, absorption spectra, 77K fluorescence spectra and resonance Raman spectra. Changes in pigment composition of the PSII RC as determined by HPLC after light treatment were as follows： with Increasing illumination time chlorophyll （Chl） a and β-carotene （β-car） content decreased. However, decreases in pheophytin （Pheo） could not be observed because of the mixture of the Pheo formed by degraded chlorophyll possibly. On the basis of absorption spectra, it was determined that, with a short time of illuminatlon, the initial bleaching occurred maximally at 680 nm but that with Increasing Illumination time there was a blue shift to 678 nm. It was suggested that P680 was destroyed Initially, followed by the accessory chlorophyll. The activity of P680 was almost lost after 10 mln light treatment. Moreover, the bleaching of Pheo and β-car was observed at the beginning of illumination. After Illumination, the fluorescence emission Intensity changed and the fluorescence maximum blue shifted, showing that energy transfer was disturbed. Resonance Raman spectra of the PSII RC excited at 488.0 and 514.5 nm showed four main bands, peaking at 1 527 cm^-1 （υ101）, 1 159 cm^-1 （υ2）, 1 006 cm^-1 （υ3）, 966 cm^-1 （υ4） for 488.0 nm excitation and 1 525 cm^-1 （υ1）, 1 159 cm^-1 （υ2）, 1 007 cm^-1 （υ3）, 968 cm^-1 （υ4） for 514.5 nm excitation. It was confirmed that two spectroscopically different β-car molecules exist In the PSII RC. After light treatment for 20 mln, band positions and bandwidths were unchanged. This indicates that carotenoid configuration Is not the parameter that regulates photoprotectlon in the PSII RC.     

Determination of the primary charge separation rate in Photosystem II reaction centers at 15 K

We have measured the rate constant for the formation of the oxidized chlorophyll a electron donor (P680⁺) and the reduced electron acceptor pheophytin a ^– (Pheo a ^–) following excitation of isolated Photosystem II reaction centers (PS II RC) at 15 K. This PS II RC complex consists of D₁, D₂, and cytochrome b-559 proteins and was prepared by a procedure which stabilizes the protein complex. Transient absorption difference spectra were measured from 450–840 nm as a function of time with 500fs resolution following 610 nm laser excitation. The formation of P680⁺-Pheo a ^– is indicated by the appearance of a band due to P680⁺ at 820 nm and corresponding absorbance changes at 490, 515 and 546 nm due to the formation of Pheo a ^–. The appearance of the 490 nm and 820 nm bands is monoexponenital with =1.4±0.2 ps. Treatment of the PS II RC with sodium dithionite and methyl viologen followed by exposure to laser excitation results in accumulation of Pheo a ^–. Laser excitation of these prereduced RCs at 15 K results in formation of a transient absorption spectrum assigned to ^1*P680. We observe wavelength-dependent kinetics for the recovery of the transient bleach of the Q_y absorption bands of the pigments in both untreated and pre-reduced PS II RCs at 15K. This result is attributed to an energy transfer process within the PS II RC at low temperature that is not connected with charge separation.Abbreviations PS I Photosystem I - PS II Photosystem II - RC reaction center - P680 primary electron donor in Photosystem II - Chl a chlorophyll a - Pheo a pheophytin a     

Pigment organization and their interactions in reaction centers of photosystem II: optical spectroscopy at 6 K of reaction centers with modified pheophytin composition

Photosystem II reaction centers (RC) with selectively exchanged pheophytin (Pheo) molecules as described in [Germano, M., Shkuropatov, A. Ya., Permentier, H., Khatypov, R. A., Shuvalov, V. A., Hoff, A. J., and van Gorkom, H. J. (2000) Photosynth. Res. 64, 189-198] were studied by low-temperature absorption, linear and circular dichroism, and triplet-minus-singlet absorption-difference spectroscopy. The ratio of extinction coefficients epsilon(Pheo)/epsilon(Chl) for Q(Y) absorption in the RC is approximately 0.40 at 6 K and approximately 0.45 at room temperature. The presence of 2 beta-carotenes, one parallel and one perpendicular to the membrane plane, is confirmed. Absorption at 670 nm is due to the perpendicular Q(Y) transitions of the two peripheral chlorophylls (Chl) and not to either Pheo. The "core" pigments, two Pheo and four Chl absorb in the 676-685 nm range. Delocalized excited states as predicted by the "multimer model" are seen in the active branch. The inactive Pheo and the nearby Chl, however, mainly contribute localized transitions at 676 and 680 nm, respectively, although large CD changes indicate that exciton interactions are present on both branches. Replacement of the active Pheo prevents triplet formation, causes an LD increase at 676 and 681 nm, a blue-shift of 680 nm absorbance, and a bleach of the 685 nm exciton band. The triplet state is mainly localized on the Chl corresponding to B(A) in purple bacteria. Both Pheo Q(Y) transitions are oriented out of the membrane plane. Their Q(X) transitions are parallel to that plane, so that the Pheos in PSII are structurally similar to their homologues in purple bacteria.     

Pheophytin-mediated energy storage of photosystem II particles detected by photoacoustic spectroscopy

The photoacoustic (PA) characteristics (energy storage and heat dissipation) of photosystem II (PSII) core-enriched particles from barley were studied (i) in conditions where there was electron flow, i.e., in the presence of a combination of the electron acceptor K₃ Fe (CN)₆, referred to as FeCN, and the electron donor diphenylcarbazide (DPC), and (ii) in conditions where electron flow was suppressed, i.e., in the absence of FeCN and DPC. The experimental data show that a decrease of heat dissipation with a minimum at 540 nm can be interpreted as energy storage resulting from the presence of pheophytin (Pheo) in the PSII particles. On account of the capability of the PA method to measure the energy absorbed by the chromophores which is converted to heat, it is suggested that the PA detection of Pheo present in the PSII complex will permit to clarify the function of processes involving non-radiative relaxation of excited states in P680-Pheo-Q_A interactions.Abbreviations -Car -Carotene - Chl Chlorophyll - DPC Diphenylcarbazide - EPR Electron Paramagnetic Resonance - FeCN potassium ferricyanide - HEPES N-2-hydroxyethylenepiperazine-N-2-ethanesulfonate - P680 reaction center of PSII - PA Photoacoustic - Pheo pheophytin - PSI photosystem I - PSII photosystem II - Q_A primary electron acceptor of PSII     

Modulating the redox potential of the stable electron acceptor, Q(B), in mutagenized photosystem II reaction centers

One of the unique features of electron transfer processes in photosystem II (PSII) reaction centers (RC) is the exclusive transfer of electrons down only one of the two parallel cofactor branches. In contrast to the RC core polypeptides (psaA and psaB) of photosystem I (PSI), where electron transfer occurs down both parallel redox-active cofactor branches, there is greater protein-cofactor asymmetry between the PSII RC core polypeptides (D1 and D2). We have focused on the identification of protein-cofactor relationships that determine the branch along which primary charge separation occurs (P(680)(+)/pheophytin(-)(Pheo)). We have previously shown that mutagenesis of the strong hydrogen-bonding residue, D1-E130, to less polar residues (D1-E130Q,H,L) shifted the midpoint potential of the Pheo(D1)/Pheo(D1)(-) couple to more negative values, reducing the quantum yield of primary charge separation. We did not observe, however, electron transfer down the inactive branch in D1-E130 mutants. The protein residue corresponding to D1-E130 on the inactive branch is D2-Q129 which presumably has a reduced hydrogen-bonding interaction with Pheo(D2) relative to the D1-E130 residue with Pheo(D1). Analysis of the recent 2.9 ? cyanobacterial PSII crystal structure indicated, however, that the D2-Q129 residue was too distant from the Pheo(D2) headgroup to serve as a possible hydrogen bond donor and directly impact its midpoint potential as well as potentially determine the directionality of electron transfer. Our objective was to characterize the function of this highly conserved inactive branch residue by replacing it with a nonconservative leucine or a conservative histidine residue. Measurements of Chl fluorescence decay kinetics and thermoluminescence studies indicate that the mutagenesis of D2-Q129 decreases the redox gap between Q(A) and Q(B) due to a lowering of the redox potential of Q(B). The resulting increased yield of S(2)Q(B)(-) charge recombination in the D2-Q129 mutants leads to an increased susceptibility to photoinhibitory light presumably due to (3)P(680)-mediated oxidative damage. The results indicate that the D2-Q129 residue plays a critical role in stabilizing the charge-separated state in PSII and further documents the structural and functional asymmetry between the two cofactor branches in PSII.     

Correlation of the light-induced change of absorbance with ESR signal of photosystem II in presence of silicomolybdate.

The light-induced dark-reversible ESR signal in chloroplast fragments enriched in photosystem II and free from P700 contamination has been observed in the presence of silicomolybdate as an electron acceptor operating directly on the photosystem II primary acceptor. The signal at g = 2.0025 and with line-width ΔHpp = 9G rises and decays in close correlation with the photobleaching band centered at 680 nm and the minor peak at 435 nm.     

The effects of light-induced reduction of the photosystem II reaction center

Accumulation of reduced pheophytin a (Pheo-D1) in photosystem II reaction center (PSII RC) under illumination at low redox potential is accompanied by changes in absorbance and circular dichroism spectra. The temperature dependences of these spectral changes have the potential to distinguish between changes caused by the excitonic interaction and temperature-dependent processes. We observed a conformational change in the PSII RC protein part and changes in the spatial positions of the PSII RC pigments of the active D1 branch upon reduction of Pheo-D1 only in the case of high temperature (298 K) dynamics. The resulting absorption difference spectra of PSII RC models equilibrated at temperatures of 77 K and 298 K were highly consistent with our previous experiments in which light-induced bleaching of the PSII RC absorbance spectrum was observable only at 298 K. These results support our previous hypothesis that Pheo-D1 does not interact excitonically with the other chlorins of the PSII RC, since the reduced form of Pheo-D1 causes absorption spectra bleaching only due to temperature-dependent processes. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.

Light-induced absorbance changes in chloroplasts mediated by Photosystem I and Photosystem II at low temperature

Stable light-induced absorbance changes in chloroplasts at −196 °C were measured across the visible spectrum from 370 to 730 nm in an effort to find previously undiscovered absorbance changes that could be related to the primary photochemical activity of Photosystem I or Photosystem II. A Photosystem I mediated absorbance increase of a band at 690 nm and a Photosystem II mediated absorbance increase of a band at 683 nm were found. The 690-nm change accompanied the oxidation of P₇₀₀ and the 683-nm increase accompanied the reduction of C-550. No Soret band was detected for P₇₀₀.

A specific effort was made to measure the difference spectrum for the photooxidation of P₆₈₀ under conditions (chloroplasts frozen to −196 °C in the presence of ferricyanide) where a stable, Photosystem II mediated EPR signal, attributed to P₆₈₀⁺ has been reported. The difference spectra, however, did not show that P₆₈₀⁺ was stable at −196 °C under any conditions tested. Absorbance measurements induced by saturating flashes at −196 °C (in the presence or absence of ferricyanide) indicated that all of the P₆₈₀⁺ formed by the flash was reduced in the dark either by a secondary electron donor or by a backreaction with the primary electron acceptor. We conclude that P₆₈₀⁺ is not stable in the dark at −196 °C: if the normal secondary donor at −196 °C is oxidized by ferricyanide prior to freezing, P₆₈₀⁺ will oxidize other substances.     

Shortening of life-time of the pair [P680+Pheo-] contributes to general inhibitory effect of dinoseb on electron transfer in PS-II

It is shown that dinoseb, added to subchloroplast photosystem-II (PS-II) preparations from pea at a concentration higher than 5 microM, along with blocking the electron transfer on the acceptor side of PS-II, induces the following effects revealing its capability to have redox interaction with the components of PS-II reaction center (RC)-pheophytin (Pheo) and chlorophyll P680: (1) acceleration of the dark relaxation of absorbance (delta A) and chlorophyll fluorescence (delta F) changes related to photoreduction of Pheo as a result of the photoreaction [P680Pheo] [symbol: see text] [P680Pheo-] that leads to elimination of the delta A and delta F at a concentration of the inhibitor higher than 50 microM; (2) lowering of the maximum level of fluorescence (F) due to a decrease of delta F under the condition when the electron acceptor, QA, is reduced; (3) loss of the described effects of dinoseb and appearance of its capability to donate electron to RC of PS-II in the presence of dithionite which reduces dinoseb in the dark; (4) inhibition of delta A related to photooxidation of P680; (5) activation of delta A related to photooxidation P700 in photosystem-I (PS-I) preparations (a similar effect is observed upon the addition of 0.2 mM methylviologen). It is suggested that redox interaction with the pair [P680+Pheo-] leading to the shortening of its life-time contributes to the general effect of inhibition of electron transfer in PS-II by dinoseb.     

Singlet oxygen production in photosystem II and related protection mechanism

High-light illumination of photosynthetic organisms stimulates the production of singlet oxygen by photosystem II (PSII) and causes photo-oxidative stress. In the PSII reaction centre, singlet oxygen is generated by the interaction of molecular oxygen with the excited triplet state of chlorophyll (Chl). The triplet Chl is formed via charge recombination of the light-induced charge pair. Changes in the midpoint potential of the primary electron donor P(680) of the primary acceptor pheophytin or of the quinone acceptor Q(A), modulate the pathway of charge recombination in PSII and influence the yield of singlet oxygen formation. The involvement of singlet oxygen in the process of photoinhibition is discussed. Singlet oxygen is efficiently quenched by beta-carotene, tocopherol or plastoquinone. If not quenched, it can trigger the up-regulation of genes, which are involved in the molecular defence response of photosynthetic organisms against photo-oxidative stress.     

CD Spectra of D1/D2/Cyt b559 Complax in Photodamaged Photosystem Ⅱ Reaction Center

The CD spectrum of photosystem Ⅱ reaction center D1/D2/Cyt b559 complex showed a strong reverse band with positive peak at 680 nm and negative peak at 660 nm in the red absorption region (Qy band). After the D1/D2/Cyt b559 complex was illuminated by strong light, the CD signals of the complex decreased significantly in the red region in which the negative peak still existed but the positive one disappeared. The result suggested that the CD signal of photosystem Ⅱ reaction center D1/D2/Cyt b559 complex not only came from the primary donor, P680, but also from other pigments such as from accessory Chl a or Pheo a.     

Effect of NADP on light-induced cytochrome changes in membrane fragments from a blue-green alga.

The effect of NADP+ on light-induced steady-state redox changes of membrane-bound cytochromes was investigated in membrane fragements prepared from the blue-green algae Nostoc muscorum (Strain 7119) that had high rates of electron transport from water to NADP+ and from an artificial electron donor, reduced dichlorophenolindophenol (DCIPH2) to NDAP+. The membrane fragments contained very little phycocyanin and had excellent optical properties for spectrophotometric assays. With DCIPH2 as the electron donor, NADP+ had no effect on the light-induced redox changes of cytochromes: with or without NADP+, 715- or 664-nm illumination resulted mainly in the oxidation of cytochrome f and of other component(s) which may include a c-type cytochrome with an alpha peak at 549nm. With 664 nm illumination and water as the electron donor, NADP+ had a pronounced effect on the redox state of cytochromes, causing a shift toward oxidation of a component with a peak at 549 nm (possibly a c-type cytochrome), cytochrome f, and particularly cytochrome b559. Cytochrome b559 appeared to be a component of the main noncyclic electron transport chain and was photooxidized at physiological temperatures by Photosystem II. This photooxidation was apparent only in the presence of a terminal acceptor (NADP+) for the electron flow from water.     

Photoconsumption of oxygen in photosystem II preparations under impairment of the water-oxidizing complex

Oxygen consumption in photosystem II (PSII) preparations in the light was 2 mol O₂/h per mg Chl at weakly acidic and at neutral pH values. It increased fourfold to fivefold at pH 8.5-9.0. The addition of either artificial electron donors for PSII such as MnCl₂ or diphenylcarbazide, or diuron as an inhibitor of electron transfer from Q_A, the primary bound quinone acceptor, to Q_B, the secondary bound quinone acceptor of PSII, resulted in a decrease in oxygen consumption rate at basic pH to value close to ones measured at pH 6.5. Such additions did not affect oxygen consumption at lower pH values. The induction of variable chlorophyll fluorescence yield in the light differed greatly at pH 6.5 and 8.5. While at pH 6.5 the fluorescence yield, after an initial fast rise almost to F_max, only slightly decreased, at pH 8.5 after such a rise it dropped promptly to a low value. The additions of the artificial electron donors at pH 8.5 resulted in the induction kinetics close to that observed at pH 6.5. These data indicate impairment of electron donation to P680⁺ that could be caused by damage to the water oxidation system at basic pH values. In experiments with PSII preparations treated with Tris to destroy the water-oxidizing complex, photoconsumption of oxygen in the entire pH region was close to the values in untreated preparations at basic pH. In untreated preparations the rate of light-induced oxygen consumption decreased in the presence of catalase, which decomposes H₂O₂, as well as in the presence of electron acceptor potassium ferricyanide. From these data it is suggested that the light-induced oxygen consumption in PSII is caused by two processes, by an interaction of O₂ with organic radicals, which were formed due to oxidation of components of the donor side of this photosystem (proteins, lipids, pigments) by cation-radical P680⁺, as well as by oxygen reduction by still unidentified components of PSII.     

Circular dichroism of the peripheral chlorophylls in photosystem II reaction centers revealed by electrochemical oxidation

Visible absorption spectra and circular dichroism (CD) of the red absorption band of isolated photosystem II reaction centers were measured at room temperature during progressive bleaching by electrochemical oxidation, in comparison with aerobic photochemical destruction, and with anaerobic photooxidation in the presence of the artificial electron acceptor silicomolybdate. Initially, selective bleaching of peripheral chlorophylls absorbing at 672 nm was obtained by electrochemical oxidation at +0.9 V, whereas little selectivity was observed at higher potentials. Illumination in the presence of silicomolybdate did not cause a bleaching but a spectral broadening of the 672-nm band was observed, apparently in response to the oxidation of carotene. The 672-nm absorption band is shown to exhibit a positive CD, which accounts for the 674-nm shoulder in CD spectra at low temperature. The origin of this CD is discussed in view of the observation that all CD disappears with the 680-nm absorption band during aerobic photodestruction.     

Circular dichroism of the peripheral chlorophylls in photosystem II reaction centers revealed by electrochemical oxidation

Visible absorption spectra and circular dichroism (CD) of the red absorption band of isolated photosystem II reaction centers were measured at room temperature during progressive bleaching by electrochemical oxidation, in comparison with aerobic photochemical destruction, and with anaerobic photooxidation in the presence of the artificial electron acceptor silicomolybdate. Initially, selective bleaching of peripheral chlorophylls absorbing at 672 nm was obtained by electrochemical oxidation at +0.9 V, whereas little selectivity was observed at higher potentials. Illumination in the presence of silicomolybdate did not cause a bleaching but a spectral broadening of the 672-nm band was observed, apparently in response to the oxidation of carotene. The 672-nm absorption band is shown to exhibit a positive CD, which accounts for the 674-nm shoulder in CD spectra at low temperature. The origin of this CD is discussed in view of the observation that all CD disappears with the 680-nm absorption band during aerobic photodestruction.