Current state of the primary charge separation mechanism in photosystem I of cyanobacteria.

This review analyzes new data on the mechanism of ultrafast reactions of primary charge separation in photosystem I (PS I) of cyanobacteria obtained in the last decade by methods of femtosecond absorption spectroscopy. Cyanobacterial PS I from many species harbours 96 chlorophyll a (Chl a) molecules, including six specialized Chls denoted Chl1A/Chl1B (dimer P700, or PAPB), Chl2A/Chl2B, and Chl3A/Chl3B arranged in two branches, which participate in electron transfer reactions. The current data indicate that the primary charge separation occurs in a symmetric exciplex, where the special pair P700 is electronically coupled to the symmetrically located monomers Chl2A and Chl2B, which can be considered together as a symmetric exciplex Chl2APAPBChl2B with the mixed excited (Chl2APAPBChl2B)* and two charge-transfer states P700 +Chl2A - and P700 +Chl2B -. The redistribution of electrons between the branches in favor of the A-branch occurs after reduction of the Chl2A and Chl2B monomers. The formation of charge-transfer states and the symmetry breaking mechanisms were clarified by measuring the electrochromic Stark shift of ß-carotene and the absorption dynamics of PS I complexes with the genetically altered Chl 2B or Chl 2A monomers. The review gives a brief description of the main methods for analyzing data obtained using femtosecond absorption spectroscopy. The energy levels of excited and charge-transfer intermediates arising in the cyanobacterial PS I are critically analyzed.

Generation of ion-radical chlorophyll states in the light-harvesting antenna and the reaction center of cyanobacterial photosystem I.

The energy and charge-transfer processes in photosystem I (PS I) complexes isolated from cyanobacteria Thermosynechococcus elongatus and Synechocystis sp. PCC 6803 were investigated by pump-to-probe femtosecond spectroscopy. The formation of charge-transfer (CT) states in excitonically coupled chlorophyll a complexes (exciplexes) was monitored by measuring the electrochromic shift of ß-carotene in the spectral range 500-510 nm. The excitation of high-energy chlorophyll in light-harvesting antenna of both species was not accompanied by immediate appearance of an electrochromic shift. In PS I from T. elongatus, the excitation of long-wavelength chlorophyll (LWC) caused a pronounced electrochromic effect at 502 nm assigned to the appearance of CT states of chlorophyll exciplexes. The formation of ion-radical pair P700+A1- at 40 ps was limited by energy transfer from LWC to the primary donor P700 and accompanied by carotenoid bleach at 498 nm. In PS I from Synechocystis 6803, the excitation at 720 nm produced an immediate bidentate bleach at 690/704 nm and synchronous carotenoid response at 508 nm. The bidentate bleach was assigned to the formation of primary ion-radical state PB+Chl2B-, where negative charge is localized predominantly at the accessory chlorophyll molecule in the branch B, Chl2B. The following decrease of carotenoid signal at ~ 5 ps was ascribed to electron transfer to the more distant molecule Chl3B. The reduction of phylloquinone in the sites A1A and A1B was accompanied by a synchronous blue-shift of the carotenoid response to 498 nm, pointing to fast redistribution of unpaired electron between two branches in favor of the state PB+A1A-.

Mutation H(M202)L does not lead to the formation of a heterodimer of the primary electron donor in reaction centers of Rhodobacter sphaeroides when combined with mutation I(M206)H.

In photosynthetic reaction centers (RCs) of purple bacteria, conserved histidine residues [His L173 and His M202 in Rhodobacter (Rba.) sphaeroides] are known to serve as fifth axial ligands to the central Mg atom of the bacteriochlorophyll (BChl) molecules (PA and PB, respectively) that constitute the homodimer (BChl/BChl) primary electron donor P. In a number of previous studies, it has been found that replacing these residues with leucine, which cannot serve as a ligand to the Mg ion of BChl, leads to the assembly of heterodimer RCs with P represented by the BChl/BPheo pair. Here, we show that a homodimer P is assembled in Rba. sphaeroides RCs if the mutation H(M202)L is combined with the mutation of isoleucine to histidine at position M206 located in the immediate vicinity of PB. The resulting mutant H(M202)L/I(M206)H RCs are characterized using pigment analysis, redox titration, and a number of spectroscopic methods. It is shown that, compared to wild-type RCs, the double mutation causes significant changes in the absorption spectrum of the P homodimer and the electronic structure of the radical cation P+, but has only minor effect on the pigment composition, the P/P+ midpoint potential, and the initial electron-transfer reaction. The results are discussed in terms of the nature of the axial ligand to the Mg of PB in mutant H(M202)L/I(M206)H RCs and the possibility of His M202 participation in the previously proposed through-bond route for electron transfer from the excited state P* to the monomeric BChl BA in wild-type RCs.

Coherent intradimer dynamics in reaction centers of photosynthetic green bacterium Chloroflexus aurantiacus.

Early-time dynamics of absorbance changes (light minus dark) in the long-wavelength Qy absorption band of bacteriochlorophyll dimer P of isolated reaction centers (RCs) from thermophilic green bacterium Chloroflexus (Cfx.) aurantiacus was studied by difference pump-probe spectroscopy with 18-fs resolution at cryogenic temperature. It was found that the stimulated emission spectrum gradually moves to the red on the ~100-fs time scale and subsequently oscillates with a major frequency of ~140 cm-1. By applying the non-secular Redfield theory and linear susceptibility theory, the coherent dynamics of the stimulated emission from the excited state of the primary electron donor, bacteriochlorophyll dimer P*, was modeled. The model showed the possibility of an extremely fast transition from the locally excited state P1* to the spectrally different excited state P2*. This transition is clearly seen in the kinetics of the stimulated emission at 880 and 945 nm, where mostly P1* and P2* states emit, respectively. These findings are similar to those obtained previously in RCs of the purple bacterium Rhodobacter (Rba.) sphaeroides. The assumption about the existence of the second excited state P2* helps to explain the complicated temporal behavior of the ΔA spectrum measured by pump-probe spectroscopy. It is interesting that, in spite of the strong coupling between the P1* and P2* states assumed in our model, the form of the coherent oscillations is mainly defined by pure vibrational coherence in the excited states. A possible nature of the P2* state is discussed.

Ultrafast excited-state dynamics in chlorosomes isolated from the photosynthetic filamentous green bacterium Chloroflexus aurantiacus.

Bacteriochlorophyll (BChl) c pigments in the aggregated state are responsible for efficient light harvesting in chlorosomes of the filamentous anoxygenic photosynthetic bacterium, Chloroflexus (Cfx.) aurantiacus. Absorption of light creates excited states in the BChl c aggregates. After subpicosecond intrachlorosomal energy transfer, redistribution and relaxation, the excitation is transferred to the BChl a complexes and further to reaction centers on the picosecond time scale. In this work, the femtosecond excited state dynamics within BChl c oligomers of isolated Cfx. aurantiacus chlorosomes was studied by double difference pump-probe spectroscopy at room temperature. Difference (Alight - Adark ) spectra corresponding to excitation at 725 nm (blue side of the BChl c absorption band) were compared with those corresponding to excitation at 750 nm (red side of the BChl c absorption band). A very fast (time constant 70 ± 10 fs) rise kinetic component was found in the stimulated emission (SE) upon excitation at 725 nm. This component was absent at 750-nm excitation. These data were explained by the dynamical red shift of the SE due to excited state relaxation. The nature and mechanisms of the ultrafast excited state dynamics in chlorosomal BChl c aggregates are discussed.

Vyacheslav (Slava) Klimov (1945-2017): A scientist par excellence, a great human being, a friend, and a Renaissance man.

Vyacheslav Vasilevich (V.V.) Klimov (or Slava, as most of us called him) was born on January 12, 1945 and passed away on May 9, 2017. He began his scientific career at the Bach Institute of Biochemistry of the USSR Academy of Sciences (Akademy Nauk (AN) SSSR), Moscow, Russia, and then, he was associated with the Institute of Photosynthesis, Pushchino, Moscow Region, for about 50 years. He worked in the field of biochemistry and biophysics of photosynthesis. He is known for his studies on the molecular organization of photosystem II (PSII). He was an eminent scientist in the field of photobiology, a well-respected professor, and, above all, an outstanding researcher. Further, he was one of the founding members of the Institute of Photosynthesis in Pushchino, Russia. To most, Slava Klimov was a great human being. He was one of the pioneers of research on the understanding of the mechanism of light energy conversion and of water oxidation in photosynthesis. Slava had many collaborations all over the world, and he is (and will be) very much missed by the scientific community and friends in Russia as well as around the World. We present here a brief biography and some comments on his research in photosynthesis. We remember him as a friendly and enthusiastic person who had an unflagging curiosity and energy to conduct outstanding research in many aspects of photosynthesis, especially that related to PSII.

Analysis of the transformation effect in cytochrome b559 of photosystem II in terms of the model of the heme-quinone redox interaction.

Transformation of three-component redox pattern of cytochrome (Cyt) b559 in PS II membrane fragments upon various treatments is manifested in decrease of the relative content (R) of the high potential (HP) redox form of Cyt b559 and concomitant increase in the fractions of the two lower potential forms. Redox titration of Cyt b559 in different types of PS II membrane preparations was performed and revealed that (1) alteration of redox titration curve of Cyt b559 upon treatment of a sample is not specific to the type of treatment; (2) each value of RHP defines the individual shape of the redox titration curve; (3) population of Cyt b559 may exist in several stable forms with multicomponent redox pattern: three types of three-component redox pattern and one type of two-component redox pattern as well as in the form with a single Em; (4) transformation of Cyt b559 proceeds as successive conversion between the stable forms with multicomponent redox pattern; (5) upon harsh treatments, Cyt b559 abruptly converts into the state with a single Em which value is intermediate between the Em values of the two lower potential forms. Analysis of the data using the model of Cyt b559-quinone redox interaction revealed that diminution of RHP in a range from 80 to 10% reflects a shift in redox equilibrium between the heme group of Cyt b559 and the interacting quinone, due to a gradual decrease of 90 mV in Em of the heme group at the virtually unchanged Em of the quinone component.

Mechanism of adiabatic primary electron transfer in photosystem I: Femtosecond spectroscopy upon excitation of reaction center in the far-red edge of the QY band.

The ultrafast primary charge separation in Photosystem I (PS I) excited by femtosecond pulses centered at 720 and 760nm was studied by pump-to-probe laser spectroscopy. The absorbance in the red edge of PS I absorption spectrum has an unusual exponential dependence on wavelength. The cutoff of short wavelength components of 760nm pulse allows direct excitation of reaction center chlorophyll molecules without involvement of light-harvesting antenna. The transient spectrum manifests the features of the primary ion-radical pair P700+A0- at time delay <180fs, followed by formation of the secondary pair P700+A1- with a characteristic time of 26ps. The obtained data are rationalized in the framework of adiabatic three-state model that includes the chlorophyll dimer P700 and two symmetrically arranged nearest chlorophyll molecules of A0. The arrangement of chlorophylls results in strong electronic coupling between P700 and A0. Excitation in the maximum of P700 absorption generates electronic states with the highest contribution from P700*, whereas excitation in the far-red edge predominantly generates charge transfer state P700+A0- in both branches of redox-cofactors. The three-level model accounts for a flat-bottomed potential surface of the excited state and adiabatic character of electron transfer between P700 and A0, providing a microscopic explanation of the ultrafast formation of P700+A0- and exponential decline of PS I absorption.

Characterization of the low-temperature triplet state of chlorophyll in photosystem II core complexes: Application of phosphorescence measurements and Fourier transform infrared spectroscopy.

Phosphorescence measurements at 77 K and light-induced FTIR difference spectroscopy at 95 K were applied to study of the triplet state of chlorophyll a ((3)Chl) in photosystem II (PSII) core complexes isolated from spinach. Using both methods, (3)Chl was observed in the core preparations with doubly reduced primary quinone acceptor QA. The spectral parameters of Chl phosphorescence resemble those in the isolated PSII reaction centers (RCs). The main spectral maximum and the lifetime of the phosphorescence corresponded to 955±1 nm and of 1.65±0.05 ms respectively; in the excitation spectrum, the absorption maxima of all core complex pigments (Chl, pheophytin a (Pheo), and ß-carotene) were observed. The differential signal at 1667(-)/1628(+)cm(-1) reflecting a downshift of the stretching frequency of the 13(1)-keto C=O group of Chl was found to dominate in the triplet-minus-singlet FTIR difference spectrum of core complexes. Based on FTIR results and literature data, it is proposed that (3)Chl is mostly localized on the accessory chlorophyll that is in triplet equilibrium with P680. Analysis of the data suggests that the Chl triplet state responsible for the phosphorescence and the FTIR difference spectrum is mainly generated due to charge recombination in the reaction center radical pair P680(+)PheoD1(-), and the energy and temporal parameters of this triplet state as well as the molecular environment and interactions of the triplet-bearing Chl molecule are similar in the PSII core complexes and isolated PSII RCs.

Towards an understanding of redox heterogeneity of the photosystem II cytochrome b559 in the native membrane.

A complex redox titration pattern of cytochrome (Cyt) b559 in preparations of thylakoid membranes and photosystem (PS) II membrane fragments is commonly attributed to the presence of three conformational forms differing by a structure of the heme microenvironment. However, despite decades of research, structural determinants underlying differences between the redox forms of Cyt b559 have not been defined. In this work, we propose a different interpretation of redox heterogeneity in the native population of Cyt b559 assuming redox interaction between the Cyt b559 heme group and a nearby bound quinone (Q). The interacting quinone is supposed to be plastoquinone QC present in the unusual singly protonated form (QCH). The model successfully explains the unique redox properties of Cyt b559 and may provide a simple and effective mechanism of redox regulation of secondary electron transport in PS II. At the present time, the model of heme-quinone redox interaction can be considered as an alternative to the idea of conformational differences between the native redox forms of Cyt b559.

Elastic Vibrations in the Photosynthetic Bacterial Reaction Center Coupled to the Primary Charge Separation: Implications from Molecular Dynamics Simulations and Stochastic Langevin Approach.

Primary electron transfer reactions in the bacterial reaction center are difficult for theoretical explication: the reaction kinetics, almost unalterable over a wide range of temperature and free energy changes, revealed oscillatory features observed initially by Shuvalov and coauthors (1997, 2002). Here the reaction mechanism was studied by molecular dynamics and analyzed within a phenomenological Langevin approach. The spectral function of polarization around the bacteriochlorophyll special pair PLPM and the dielectric response upon the formation of PL(+)PM(-) dipole within the special pair were calculated. The system response was approximated by Langevin oscillators; the respective frequencies, friction, and energy coupling coefficients were determined. The protein dynamics around PL and PM were distinctly asymmetric. The polarization around PL included slow modes with the frequency 30-80 cm(-1) and the total amplitude of 130 mV. Two main low-frequency modes of protein response around PM had frequencies of 95 and 155 cm(-1) and the total amplitude of 30 mV. In addition, a slowly damping mode with the frequency of 118 cm(-1) and the damping time >1.1 ps was coupled to the formation of PL(+)PM(-) dipole. It was attributed to elastic vibrations of α-helices in the vicinity of PLPM. The proposed trapping of P excitation energy in the form of the elastic vibrations can rationalize the observed properties of the primary electron transfer reactions, namely, the unusual temperature and ΔG dependences, the oscillating phenomena in kinetics, and the asymmetry of the charge separation reactions.

Low-temperature (77 K) phosphorescence of triplet chlorophyll in isolated reaction centers of photosystem II.

Phosphorescence characterized by the main emission band at 952 ± 1 nm (1.30 eV), the lifetime of 1.5 ± 0.1 ms and the quantum yield nearly equal to that for monomeric chlorophyll a in aqueous detergent dispersions, has been detected in isolated reaction centers (RCs) of spinach photosystem II at 77 K. The excitation spectrum shows maxima corresponding to absorption bands of chlorophyll a, pheophytin a, and ß-carotene. The phosphorescence intensity strongly depends upon the redox state of RCs. The data suggest that the phosphorescence signal originates from the chlorophyll triplet state populated via charge recombination in the radical pair [Formula: see text].

My journey in photosynthesis research.

At the invitation of Suleyman I. Allakhverdiev, I provide here a brief autobiography for this special issue that recognizes my service and research for the larger international community of photosynthesis research.

The L(M196)H mutation in Rhodobacter sphaeroides reaction center results in new electrostatic interactions.

New histidine residue was introduced in M196 position in the reaction center of Rhodobacter sphaeroides in order to alter polarity of the BChl dimer's protein environment and to study how it affects properties and structure of the primary electron donor P. It was shown that in the absorption spectrum of the mutant RC the 6 nm red shift of the Q Y P band was observed together with considerable decrease of its amplitude. The mid-point potential of P/P (+) in the mutant RC was increased by +65 (±15) mV as compared to the E m P/P (+) value in the wild-type RC suggesting that the mutation resulted in new pigment-protein interactions. Crystal structure of RC L(M196)H determined at 2.4 Å resolution implies that BChl Р В and introduced histidine-M196 organize new electrostatic contact that may be specified either as π-π staking or as hydrogen-π interaction. Besides, the structure of the mutants RC shows that His-M196 apparently became involved in hydrogen bond network existing in BChl Р В vicinity that may favor stability of the mutant RC.

Spectral exhibition of electron-vibrational relaxation in P* state of Rhodobacter sphaeroides reaction centers.

Electron-vibrational relaxation in the excited state of the primary electron donor, bacteriochlorophyll dimer P, in the reaction centers (RCs) of purple photosynthetic bacteria Rhodobacter sphaeroides is modeled. A multimode model of three states (i.e., the ground state Pg, initially excited P1*, and relaxed excited P2*) is used to calculate the incoherent dynamics of the difference (ΔA) spectra on a femtosecond timescale for the YM210 W mutant RCs. The relaxation processes are described by the step-ladder model. The model shows that the electron-vibrational relaxation in the excited state of P is visualized by the transient red shift of the stimulated emission from P*. The dynamics of this shift is observed as a change in the ΔA spectrum shape in its red-most part, within a few hundreds of femtoseconds after excitation. As a result, an initial rise in the red-side ΔA kinetics is delayed with respect to the blue-side kinetics. The time constant of the P1* â†’ P2* electronic relaxation (54 fs) and the Pg, P1*, and P2* vibrational relaxations (120 fs), used in the model, provided the best fit of the experimental time-resolved ΔA spectra and kinetics at 90 and 293 K. The possible nature of the P1* â†’ P2* electronic relaxation is discussed.

Chemically modified reaction centers of photosystem II: Exchange of pheophytin a with 7-deformyl-7-hydroxymethyl-pheophytin b.

The native pheophytin a (Pheo a) in isolated reaction centers of photosystem II (PSII RCs) has been chemically exchanged with extraneous 7-deformyl-7-hydroxymethyl-Pheo b (7(1)-OH-Pheo b) which differs from Pheo a by the C-7 substituent (hydroxymethyl instead of methyl). The two pigments have similar reduction potentials in vitro [M. Meyer, Dissertation, Universität München, 1997], while their absorption spectra show small but distinct differences in the visible region. The resulting 7(1)-OH-Pheo b-modified reaction center preparations were characterized by high-performance liquid chromatography, electronic absorption and light-induced Fourier transform infra red absorption difference spectroscopies, together with photoaccumulation of the reduced pheophytin electron acceptor and NaBH4-treatment. About 70% of the total Pheo a molecules are found to be replaced by 7(1)-OH-Pheo b molecules in modified preparations, indicating that both the photochemically active (PheoD1) and inactive (PheoD2) binding sites were subjected to pigment exchange. The 7(1)-OH-Pheo b molecule located at the PheoD1 site is able to functionally replace the native Pheo a, participating in primary charge separation as an electron acceptor. The Qx absorption band of this modified pheophytin molecule is localized at ~546nm; its Qy band is blue-shifted with respect to the absorption of other reaction center core pigments, being located at ~665nm. The Qy and Qx optical transitions of the 7(1)-OH-Pheo b molecule exchanged into the PheoD2 site are identified at 677 and 543.5nm, respectively. The photochemically active double-modified PSII RCs additionally containing 7-deformyl-7-hydroxymethyl-13(1)-deoxo-13(1)-hydroxy-Pheo b at the PheoD2 site were obtained by treatment of the 7(1)-OH-Pheo b-modified RCs with NaBH4.

Biphasic reduction of cytochrome b559 by plastoquinol in photosystem II membrane fragments: evidence for two types of cytochrome b559/plastoquinone redox equilibria.

In photosystem II membrane fragments with oxidized cytochrome (Cyt) b559 reduction of Cyt b559 by plastoquinol formed in the membrane pool under illumination and by exogenous decylplastoquinol added in the dark was studied. Reduction of oxidized Cyt b559 by plastoquinols proceeds biphasically comprising a fast component with a rate constant higher than (10s)(-1), named phase I, followed by a slower dark reaction with a rate constant of (2.7min)(-1) at pH6.5, termed phase II. The extents of both components of Cyt b559 reduction increased with increasing concentrations of the quinols, with that, maximally a half of oxidized Cyt b559 can be photoreduced or chemically reduced in phase I at pH6.5. The photosystem II herbicide dinoseb but not 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) competed with the quinol reductant in phase I. The results reveal that the two components of the Cyt b559 redox reaction reflect two redox equilibria attaining in different time domains. One-electron redox equilibrium between oxidized Cyt b559 and the photosystem II-bound plastoquinol is established in phase I of Cyt b559 reduction. Phase II is attributed to equilibration of Cyt b559 redox forms with the quinone pool. The quinone site involved in phase I of Cyt b559 reduction is considered to be the site regulating the redox potential of Cyt b559 which can accommodate quinone, semiquinone and quinol forms. The properties of this site designated here as QD clearly suggest that it is distinct from the site QC found in the photosystem II crystal structure.

FTIR spectroscopy of the reaction center of Chloroflexus aurantiacus: photoreduction of the bacteriopheophytin electron acceptor.

Mid-infrared spectral changes associated with the photoreduction of the bacteriopheophytin electron acceptor H(A) in reaction centers (RCs) of the filamentous anoxygenic phototrophic bacterium Chloroflexus (Cfl.) aurantiacus are examined by light-induced Fourier transform infrared (FTIR) spectroscopy. The light-induced H(A)(-)/H(A) FTIR (1800-1200cm(-1)) difference spectrum of Cfl. aurantiacus RCs is compared to that of the previously well characterized purple bacterium Rhodobacter (Rba.) sphaeroides RCs. The most notable feature is that the large negative IR band at 1674cm(-1) in Rba. sphaeroides R-26, attributable to the loss of the absorption of the 13(1)-keto carbonyl of H(A) upon the radical anion H(A)(-) formation, exhibits only a very minor upshift to 1675cm(-1) in Cfl. aurantiacus. In contrast, the absorption band of the 13¹-keto C=O of H(A)(-) is strongly upshifted in the spectrum of Cfl. aurantiacus compared to that of Rba. sphaeroides (from 1588 to 1623cm(-1)). The data are discussed in terms of: (i) replacing the glutamic acid at L104 in Rba. sphaeroides R-26 RCs by a weaker hydrogen bond donor, a glutamine, at the equivalent position L143 in Cfl. aurantiacus RCs; (ii) a strengthening of the hydrogen-bonding interaction of the 13¹-keto C=O of H(A) with Glu L104 and Gln L143 upon H(A)(-) formation and (iii) a possible influence of the protein dielectric environment on the 13¹-keto C=O stretching frequency of neutral H(A). A conformational heterogeneity of the 13³-ester C=O group of H(A) is detected for Cfl. aurantiacus RCs similar to what has been previously described for purple bacterial RCs.

Femtosecond primary charge separation in Synechocystis sp. PCC 6803 photosystem I.

The ultrafast (<100 fs) conversion of delocalized exciton into charge-separated state between the primary donor P700 (bleaching at 705 nm) and the primary acceptor A0 (bleaching at 690 nm) in photosystem I (PS I) complexes from Synechocystis sp. PCC 6803 was observed. The data were obtained by application of pump-probe technique with 20-fs low-energy pump pulses centered at 720 nm. The earliest absorbance changes (close to zero delay) with a bleaching at 690 nm are similar to the product of the absorption spectrum of PS I complex and the laser pulse spectrum, which represents the efficiency spectrum of the light absorption by PS I upon femtosecond excitation centered at 720 nm. During the first approximately 60 fs the energy transfer from the chlorophyll (Chl) species bleaching at 690 nm to the Chl bleaching at 705 nm occurs, resulting in almost equal bleaching of the two forms with the formation of delocalized exciton between 690-nm and 705-nm Chls. Within the next approximately 40 fs the formation of a new broad band centered at approximately 660 nm (attributed to the appearance of Chl anion radical) is observed. This band decays with time constant simultaneously with an electron transfer to A1 (phylloquinone). The subtraction of kinetic difference absorption spectra of the closed (state P700+A0A1) PS I reaction center (RC) from that of the open (state P700A0A1) RC reveals the pure spectrum of the P700+A0- ion-radical pair. The experimental data were analyzed using a simple kinetic scheme: An*-->k1[(PA0)*A1--><100 fs P+A0-A1]-->k2P+A0A1-, and a global fitting procedure based on the singular value decomposition analysis. The calculated kinetics of transitions between intermediate states and their spectra were similar to the kinetics recorded at 694 and 705 nm and the experimental spectra obtained by subtraction of the spectra of closed RCs from the spectra of open RCs. As a result, we found that the main events in RCs of PS I under our experimental conditions include very fast (<100 fs) charge separation with the formation of the P700+A0-A1 state in approximately one half of the RCs, the approximately 5-ps energy transfer from antenna Chl* to P700A0A1 in the remaining RCs, and approximately 25-ps formation of the secondary radical pair P700+A0A1-.

Primary light-energy conversion in tetrameric chlorophyll structure of photosystem II and bacterial reaction centers: I. A review.

The purpose of the review is to show that the tetrameric (bacterio)chlorophyll ((B)Chl) structures in reaction centers of photosystem II (PSII) of green plants and in bacterial reaction centers (BRCs) are similar and play a key role in the primary charge separation. The Stark effect measurements on PSII reaction centers have revealed an increased dipole moment for the transition at approximately 730 nm (Frese et al., Biochemistry 42:9205-9213, 2003). It was found (Heber and Shuvalov, Photosynth Res 84:84-91, 2005) that two fluorescent bands at 685 and 720 nm are observed in different organisms. These two forms are registered in the action spectrum of Q(A) photoreduction. Similar results were obtained in core complexes of PSII at low temperature (Hughes et al., Biochim Biophys Acta 1757: 841-851, 2006). In all cases the far-red absorption and emission can be interpreted as indication of the state with charge transfer character in which the chlorophyll monomer plays a role of an electron donor. The role of bacteriochlorophyll monomers (B(A) and B(B)) in BRCs can be revealed by different mutations of axial ligand for Mg central atoms. RCs with substitution of histidine L153 by tyrosine or leucine and of histidine M182 by leucine (double mutant) are not stable in isolated state. They were studied in antennaless membrane by different kinds of spectroscopy including one with femtosecond time resolution. It was found that the single mutation (L153HY) was accompanied by disappearance of B(A) molecule absorption near 802 nm and by 14-fold decrease of photochemical activity measured with ms time resolution. The lifetime of P(870)* increased up to approximately 200 ps in agreement with very low rate of the electron transfer to A-branch. In the double mutant L153HY + M182HL, the B(A) appears to be lost and B(B) is replaced by bacteriopheophytin Phi(B) with the absence of any absorption near 800 nm. Femtosecond measurements have revealed the electron transfer to B-branch with a time constant of approximately 2 ps. These results are discussed in terms of obligatory role of B(A) and Phi(B) molecules located near P for efficient electron transfer from P*.