The PSII calcium site revisited.

Oxidation of H2O by photosystem II is a unique redox reaction in that it requires Ca2+ as well as Cl- as obligatory activators/cofactors of the reaction, which is catalyzed by Mn atoms. The properties of the binding site for Ca2+ in this reaction resemble those of other Ca2+ binding proteins, and recent X-ray structural data confirm that the metal is in fact ligated at least in part by amino acid side chain oxo anions. Removal of Ca2+ blocks water oxidation chemistry at an early stage in the cycle of redox reactions that result in O-O bond formation, and the intimate involvement of Ca2+ in this reaction that is implied by this result is confirmed by an ever-improving set of crystal structures of the cyanobacterial enzyme. Here, we revisit the photosystem II Ca2+ site, in part to discuss the additional information that has appeared since our earlier review of this subject (van Gorkom HJ, Yocum CF In: Wydrzynski TJ, Satoh K (eds) Photosystem II: the light-driven water:plastoquinone oxidoreductase), and also to reexamine earlier data, which lead us to conclude that all S-state transitions require Ca2+.

Energy and electron transfer in photosystem II reaction centers with modified pheophytin composition.

Energy and electron transfer in Photosystem II reaction centers in which the photochemically inactive pheophytin had been replaced by 13(1)-deoxo-13(1)-hydroxy pheophytin were studied by femtosecond transient absorption-difference spectroscopy at 77 K and compared to the dynamics in untreated reaction center preparations. Spectral changes induced by 683-nm excitation were recorded both in the Q(Y) and in the Q(X) absorption regions. The data could be described by a biphasic charge separation. In untreated reaction centers the major component had a time constant of 3.1 ps and the minor component 33 ps. After exchange, time constants of 0.8 and 22 ps were observed. The acceleration of the fast phase is attributed in part to the redistribution of electronic transitions of the six central chlorin pigments induced by replacement of the inactive pheophytin. In the modified reaction centers, excitation of the lowest energy Q(Y) transition produces an excited state that appears to be localized mainly on the accessory chlorophyll in the active branch (B(A) in bacterial terms) and partially on the active pheophytin H(A). This state equilibrates in 0.8 ps with the radical pair. B(A) is proposed to act as the primary electron donor also in untreated reaction centers. The 22-ps (pheophytin-exchanged) or 33-ps (untreated) component may be due to equilibration with the secondary radical pair. Its acceleration by H(B) exchange is attributed to a faster reverse electron transfer from B(A) to. After exchange both and are nearly isoenergetic with the excited state.

Kinetics of electron transfer from Q(a) to Q(b) in photosystem II.

The oxidation kinetics of the reduced photosystem II electron acceptor Q(A)(-) was investigated by measurement of the chlorophyll fluorescence yield transients on illumination of dark-adapted spinach chloroplasts by a series of saturating flashes. Q(A)(-) oxidation depends on the occupancy of the "Q(B) binding site", where this reaction reduces plastoquinone to plastoquinol in two successive photoreactions. The intermediate, one-electron-reduced plastosemiquinone anion Q(B)(-) remains tightly bound, and its reduction by Q(A)(-) may proceed with simple first-order kinetics. The next photoreaction, in contrast, may find the Q(B) binding site occupied by a plastoquinone, a plastoquinol, or neither of the two, resulting in heterogeneous Q(A)(-) oxidation kinetics. The assumption of monophasic Q(B)(-) reduction kinetics is shown to allow unambiguous decomposition of the observed multiphasic Q(A)(-) oxidation. At pH 6.5 the time constant for Q(A)(-) oxidation was found to be 0.2-0.4 ms with Q(B) in the site, 0.6-0.8 ms with Q(B)(-) in the site, 2-3 ms when the site is empty and Q(B) has to bind first, and of the order of 0.1 s if the site is temporarily blocked by the presence of Q(B)H(2) or other low-affinity inhibitors such as carbonyl cyanide m-chlorophenylhydrazone (CCCP). Effects of pH and H(2)O/D(2)O exchange were found to be remarkably nonspecific. No influence of the S-states could be demonstrated.

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.

Ultrahigh field MAS NMR dipolar correlation spectroscopy of the histidine residues in light-harvesting complex II from photosynthetic bacteria reveals partial internal charge transfer in the B850/His complex.

Low-temperature 15N and 13C CP/MAS (cross-polarization/magic angle spinning) NMR has been used to analyze BChl-histidine interactions and the electronic structure of histidine residues in the light-harvesting complex II (LH2) of Rhodopseudomonas acidophila. The histidines were selectively labeled at both or one of the two nitrogen sites of the imidazole ring. The resonances of histidine nitrogens that are interacting with B850 BChl a have been assigned. Specific 15N labeling confirmed that it is the tau-nitrogen of histidines which is ligated to Mg2+ of B850 BChl molecules (beta-His30, alpha-His31). The pi-nitrogens of these Mg2+-bound histidines were found to be protonated and may be involved in hydrogen bond interactions. Comparison of the 2-D MAS NMR homonuclear (13C-13C) dipolar correlation spectrum of [13C6,15N3]-histidines in the LH2 complex with model systems in the solid state reveals two different classes of electronic structures from the histidines in the LH2. In terms of the 13C isotropic shifts, one corresponds to the neutral form of histidine and the other resembles a positively charged histidine species. 15N-13C double-CP/MAS NMR data provide evidence that the electronic structure of the histidines in the neutral BChl a/His complexes resembles the positive charge character form. While the Mg...15N isotropic shift confirms a partial positive charge transfer, its anisotropy is essentially of the lone pair type. This provides evidence that the hybridization structure corresponding to the neutral form of the imidazole is capable of "buffering" a significant amount of positive charge.

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).

Photochemically induced nuclear spin polarization in reaction centers of photosystem II observed by 13C-solid-state NMR reveals a strongly asymmetric electronic structure of the P680(.+) primary donor chlorophyll.

We report (13)C magic angle spinning NMR observation of photochemically induced dynamic nuclear spin polarization (photo-CIDNP) in the reaction center (RC) of photosystem II (PS2). The light-enhanced NMR signals of the natural abundance (13)C provide information on the electronic structure of the primary electron donor P(680) (chlorophyll a molecules absorbing around 680 nm) and on the p(z) spin density pattern in its oxidized form, P(680)(.+). Most centerband signals can be attributed to a single chlorophyll a (Chl a) cofactor that has little interaction with other pigments. The chemical shift anisotropy of the most intense signals is characteristic for aromatic carbon atoms. The data reveal a pronounced asymmetry of the electronic spin density distribution within the P(680)(.+). PS2 shows only a single broad and intense emissive signal, which is assigned to both the C-10 and C-15 methine carbon atoms. The spin density appears shifted toward ring III. This shift is remarkable, because, for monomeric Chl a radical cations in solution, the region of highest spin density is around ring II. It leads to a first hypothesis as to how the planet can provide itself with the chemical potential to split water and generate an oxygen atmosphere using the Chl a macroaromatic cycle. A local electrostatic field close to ring III can polarize the electronic charge and associated spin density and increase the redox potential of P(680) by stabilizing the highest occupied molecular orbital, without a major change of color. This field could be produced, e.g., by protonation of the keto group of ring V. Finally, the radical cation electronic structure in PS2 is different from that in the bacterial RC, which shows at least four emissive centerbands, indicating a symmetric spin density distribution over the entire bacteriochlorophyll macrocycle.

Exploring the calcium-binding site in photosystem II membranes by solid-state (113)Cd NMR.

Calcium (Ca(2+)) is an essential cofactor for photosynthetic oxygen evolution. Although the involvement of Ca(2+) at the oxidizing side of photosystem II of plants has been known for a long time, its ligand interactions and mode of action have remained unclear. In the study presented here, (113)Cd magic-angle spinning solid-state NMR spectroscopy is used to probe the Ca(2+)-binding site in the water-oxidizing complex of (113)Cd(2+)-substituted PS2. A single NMR signal 142 ppm downfield from Cd(ClO(4))(2).2H(2)O was recorded from Cd(2+) present at the Ca(2+)-binding site. The anisotropy of the signal is small, as indicated by the absence of spinning side bands. The signal intensity is at its maximum at a temperature of -60 degrees C. The line width of the proton signal in a WISE (wide-line separation) two-dimensional (1)H-(113)Cd NMR experiment demonstrates that the signal arises from Cd(2+) in a solid and magnetically undisturbed environment. The chemical shift, the small anisotropy, and the narrow line of the (113)Cd NMR signal provide convincing evidence for a 6-fold coordination, which is achieved partially by oxygen and partially by nitrogen or chlorine atoms in otherwise a symmetric octahedral environment. The absence of a (113)Cd signal below -70 degrees C suggests that the Ca(2+)-binding site is close enough to the tetramanganese cluster to be affected by its electron spin state. To our knowledge, this is the first report for the application of solid-state NMR in the study of the membrane-bound PS2 protein complex.

Selective replacement of the active and inactive pheophytin in reaction centres of Photosystem II by 13(1)-deoxo-13(1)-hydroxy-pheophytin a and comparison of their 6 K absorption spectra.

Pheophytin a (Pheo) in Photosystem II reaction centres was exchanged for 13(1)-deoxo-13(1)-hydroxy-pheophytin a (13(1)-OH-Pheo). The absorption bands of 13(1)-OH-Pheo are blue-shifted and well separated from those of Pheo. Two kinds of modified reaction centre preparations can be obtained by applying the exchange procedure once (RC(1x)) or twice (RC(2x)). HPLC analysis and Pheo Q(X) absorption at 543 nm show that in RC(1x) about 50% of Pheo is replaced and in RC(2x) about 75%. Otherwise, the pigment and protein composition are not modified. Fluorescence emission and excitation spectra show quantitative excitation transfer from the new pigment to the emitting chlorophylls. Photoaccumulation of Pheo(-) is unmodified in RC(1x) and decreased only in RC(2x), suggesting that the first exchange replaces the inactive and the second the active Pheo. Comparing the effects of the first and the second replacement on the absorption spectrum at 6 K did not reveal substantial spectral differences between the active and inactive Pheo. In both cases, the absorption changes in the Q(Y) region can be interpreted as a combination of a blue shift of a transition at 684 nm, a partial decoupling of chlorophylls absorbing at 680 nm and a disappearance of Pheo absorption in the 676-680 nm region. No absorption decrease is observed at 670 nm for RC(1x) or RC(2x), showing that neither of the two reaction centre pheophytins contributes substantially to the absorption at this wavelength.

Activating anions that replace Cl- in the O2-evolving complex of photosystem II slow the kinetics of the terminal step in water oxidation and destabilize the S2 and S3 states.

Photosystem II, the multisubunit protein complex that oxidizes water to O2, requires the inorganic cofactors Ca2+ and Cl- to exhibit optimal activity. Chloride can be replaced functionally by a small number of anionic cofactors (Br-, NO3-, NO2-, I-), but among these anions, only Br- is capable of restoring rates of oxygen evolution comparable to those observed with Cl-. UV absorption difference spectroscopy was utilized in the experiments described here as a probe to monitor donor side reactions in photosystem II in the presence of Cl- or surrogate anions. The rate of the final step of the water oxidation cycle was found to depend on the activating anion bound at the Cl- site, but the kinetics of this step did not limit the light-saturated rate of oxygen evolution. Instead, the lower oxygen evolution rates supported by surrogate anions appeared to be correlated with an instability of the higher oxidation states of the oxygen-evolving complex that was induced by addition of these anions. Reduction of these states takes place not only with I- but also with NO2- and to a lesser extent even with NO3- and Br- and is not related to the ability of these anions to bind at the Cl- binding site. Rather, it appears that these anions can attack higher oxidation states of the oxygen evolving complex from a second site that is not shielded by the extrinsic 17 and 23 kDa polypeptides and cause a one-electron reduction. The decrease of the oxygen evolution rate may result from accumulated damage to the reaction center protein by the one-electron oxidation product of the anion.

S-state dependence of chloride binding affinities and exchange dynamics in the intact and polypeptide-depleted O2 evolving complex of photosystem II.

The Cl- binding properties in the successive oxidation states of the O2 evolving complex of photosystem II were investigated by measurements of UV absorbance changes, induced by a series of saturating flashes, that monitor manganese oxidation state transitions. In dark-adapted, intact photosystem II, Cl- can be replaced by NO3- in minutes, in an exchange reaction that depends on the NO3- concentration and that is not rate-limited by dissociation of Cl- from its binding site. Preillumination of dark-adapted photosystem II by one or two flashes accelerated the NO3- substitution reaction by an order of magnitude. A quantitative analysis of the Cl- concentration dependence of UV absorbance changes, measured in photosystem II preparations depleted of extrinsic 17 and 23 kDa polypeptides, shows that the Cl- binding properties of photosystem II change with the oxidation state of the oxygen evolving complex. Although the affinity for the individual S-states could not be determined with precision, it is shown that the affinity is an order of magnitude lower in the S2 state than in the S1 state. Comparison of the results obtained using intact photosystem II and preparations depleted of the 17 and 23 kDa extrinsic polypeptides suggests that these proteins constitute a diffusion barrier, which prevents fast equilibration of the Cl- binding site with the medium, but does not change the Cl- affinity of the binding site.

The photosynthetic oxygen evolving complex requires chloride for its redox state S2-->S3 and S3-->S0 transitions but not for S0-->S1 or S1-->S2 transitions.

The Cl- requirement in the redox cycle of the oxygen-evolving complex (OEC) was determined by measurements of flash-induced UV absorbance changes in Cl(-)-depleted and Cl(-)-reconstituted photosystem II membranes. On the first flash after dark adaptation the spectrum and amplitude of those changes, known to reflect the oxidation of MnIII to MnIV on the S1-->S2 transition, were the same in the presence or absence of Cl-. On the second and later flashes, however, absorbance changes in Cl(-)-depleted samples revealed only electron transfer from tyrosine to quinone which reversed slowly in the dark by charge recombination and did not produce the S3-state. A rapid method was developed to remove Cl- after producing the S3-state by two flashes. The lifetime of the S3-state was found to be unaffected by Cl(-)-depletion, in contrast to the 20-fold stabilization of the S2 lifetime by Cl- removal, and the Cl(-)-depleted S3-state did not proceed to S0 on flash illumination. However, when the same Cl(-)-depletion procedure was applied after producing the S0-state by three flashes, further advance to S2 by two additional flashes was not impaired by the absence of Cl-. The requirement for Cl- only on the S2-->S3 and S3-->S0 transitions can be rationalized by the hypothesis that Cl- is required for electron transfer between manganese ions within the oxygen-evolving complex.

Bicarbonate may Be required for ligation of manganese in the oxygen-evolving complex of photosystem II.

It was previously shown in the photosystem II membrane preparation DT-20 that photoxidation of the oxygen-evolving manganese cluster was blocked by 0.1 mM formate, unless 0.2 mM bicarbonate was present as well [Wincencjusz, H., Allakhverdiev, S. I., Klimov, V. V., and Van Gorkom, H. J. (1996) Biochim. Biophys. Acta 1273, 1-3]. Here it is shown by measurements of EPR signal II that oxidation of the secondary electron donor, YZ, is not inhibited. However, the reduction of is greatly slowed and occurs largely by back reaction with reduced acceptors. Bicarbonate is shown to prevent the loss of fast electron donation to . The release of about one or two free Mn2+ per photosystem II during formate treatment, and the fact that these effects are mimicked by Mn-depletion, suggests that formate may act by replacing a bicarbonate which is essential for Mn binding. Irreversible light-induced rebinding in an EPR-silent form of Mn2+ that was added to Mn-depleted DT-20 was indeed found to depend on the presence of bicarbonate, as did the reconstitution in such material of both the fast electron donation to and the UV absorbance changes characteristic of a functional oxygen-evolving complex. It is concluded that bicarbonate may be an essential ligand of the functional Mn cluster.

The association of different detergents with the photosynthetic reaction center protein of Rhodobacter sphaeroides R26 and the effects on its photochemistry.

Detergent-free reaction centers from Rhodobacter sphaeroides R26 were used to study the solubilization of reaction centers in various detergents and their effects on reaction center photochemistry. 500 +/- 100 n-octyl-beta-D-glucopyranoside or 51 +/- 5 Triton X-100 molecules were associated with one reaction center. For N.N-alkylamine N-oxide detergents with chain lengths in the range from 8-13 carbon atoms, the number of detergent molecules associated with the reaction centers increased with decreasing chain length. The amount of detergent molecules associated with the reaction centers decreased almost tenfold if the pH was increased from pH 6 to pH 10. The addition of 5% 1,2,3-heptanetriol to various detergents lowered the detergent/reaction center ratio by a factor of two compared to that for the pure detergent. The detergent concentration at which solubilization of the reaction center occurs was close to the critical micelle concentration for all detergents studied. The absorption band at 865 nm of the primary donor in the reaction center shifts to 846 nm when detergent was removed from the reaction center; upon resolubilization with various detergents, this band shifts back to 865 nm. In 80-90% of the detergent-free reaction centers, the secondary electron transfer from QA to QB was inhibited: this electron transfer was restored after re-addition of detergent.

Electroluminescence.

An overview is presented of research based on the observation by Arnold and Azzi (1971) (Photochem Photobiol 14: 233-240), that an electric field induces charge-recombination luminescence in a suspension of photosynthetic membrane vesicles. The 'electroluminescence' signals from Photosystems I and II are discussed in relation to the shape of the vesicles and the membrane potentials generated by the externally applied electric field. The use of the electroluminescence amplitude as a probe to study the kinetics and energetics of charge separation, and of its kinetics to monitor the electric-field induced charge recombination process are reviewed. Currently unresolved issues regarding the emission yield of electroluminescence are briefly discussed and the properties are summarized of the unexplained Photosystem II luminescence which is not sensitive to the membrane potential.

Effect of the redox state of QB on electric field-induced charge recombination in Photosystem II.

Electric field-induced charge recombination in Photosystem II (PS II) was studied in osmotically swollen spinach chloroplasts ('blebs') by measurement of the concomitant chlorophyll luminescence emission (electroluminescence). A pronounced dependence on the redox state of the two-electron gate QB was observed and the earlier failure to detect it is explained. The influence of the QB/QB (-) oscillation on electroluminescence was dependent on the redox state of the oxygen evolving complex; at times around one millisecond after flash illumination a large effect was observed in the states S2 and S3, but not in the state 'S4' (actually Z(+)S3). The presence of the oxidized secondary electron donor, tyrosine Z(+), appeared to prevent expression of the QB/QB (-) effect on electroluminescence, possibly because this effect is primarily due to a shift of the redox equilibrium between Z/Z(+) and the oxygen evolving complex.

Flash-induced redox changes in oxygen-evolving spinach Photosystem II core particles.

Flash-induced redox reactions in spinach PS II core particles were investigated with absorbance difference spectroscopy in the UV-region and EPR spectroscopy. In the absence of artificial electron acceptors, electron transport was limited to a single turnover. Addition of the electron acceptors DCBQ and ferricyanide restored the characteristic period-four oscillation in the UV absorbance associated with the S-state cycle, but not the period-two oscillation indicative of the alternating appearance and disappearance of a semiquinone at the QB-site. In contrast to PS II membranes, all active centers were in state S1 after dark adaptation. The absorbance increase associated with the S-state transitions on the first two flashes, attributed to the Z(+)S1→ZS2 and Z(+)S2→ZS3 transitions, respectively, had half-times of 95 and 380 µs, similar to those reported for PS II membrane fragments. The decrease due to the Z(+)S3→ZS0 transition on the third flash had a half-time of 4.5 ms, as in salt-washed PS II membrane fragments. On the fourth flash a small, unresolved, increase of less than 3 µs was observed, which might be due to the Z(+)S0→ZS1 transition. The deactivation of the higher S-states was unusually fast and occurred within a few seconds and so was the oxidation of S0 to S1 in the dark, which had a half-time of 2-3 min. The same lifetime was found for tyrosine D(+), which appeared to be formed within milliseconds after the first flash in about 10% inactive centers and after the third and later flashes by active centers in Z(+)S3.

Kok's oxygen clock: What makes it tick? The structure of P680 and consequences of its oxidizing power.

New insights in the structure of P680, the primary electron donor in Photosystem II, are summarized and the implications of its oxidizing power for energy transfer and singlet oxygen production are discussed.

Absorbance difference spectra of the S-state transitions in Photosystem II core particles.

Redox changes of the oxygen evolving complex in PS II core particles were investigated by absorbance difference spectroscopy in the UV-region. The oscillation of the absorbance changes induced by a series of saturating flashes could not be explained by the minimal Kok model (Kok et al. 1970) consisting of a 4-step redox cycle, S0 → S1 → S2 → S3 → S0, although the values of most of the relevant parameters had been determined experimentally. Additional assumptions which allow a consistent fit of all data are a slow equilibration of the S3 state with an inactive state, perhaps related to Ca(2+)-release, and a low quantum efficiency for the first turnover after dark-adaptation. Difference spectra of the successive S-state transitions were determined. At wavelengths above 370 nm, they were very different due to the different contribution of a Chl bandshift in each spectrum. At shorter wavelengths, the S1 → S2 transition showed a difference spectrum similar to that reported by Dekker et al. 1984b and attributed to an Mn(III) to Mn(IV) oxidation. The spectrum of absorbance changes associated with the S2 → S3 transition was similar to that reported by Lavergne 1991 for PS II membranes. The S0 → S1 transition was associated with a smaller but still substantial absorbance increase in the UV. Differences with the spectra reported by Lavergne 1991 are attributed to electrostatic effects on electron transfer at the acceptor side associated with the S-state dependence of proton release in PS II membranes.

Rapid and simple isolation of pure photosystem II core and reaction center particles from spinach.

Pure and active oxygen-evolving PS II core particles containing 35 Chl per reaction center were isolated with 75% yield from spinach PS II membrane fragments by incubation with n-dodecyl-ß-D-maltoside and a rapid one step anion-exchange separation. By Triton X-100 treatment on the column these particles could be converted with 55% yield to pure and active PS II reaction center particles, which contained 6 Chl per reaction center.