V. V. Gorokhov

Photophysical properties of carborane-containing derivatives of 5,10,15,20-tetra(p-aminophenyl)porphyrin

Absorption, fluorescence emission, fluorescence excitation spectra and fluorescence decay kinetics of carborane derivatives of 5,10,15,20-tetra(p-aminophenyl)porphyrin have been investigated. Carborane derivatives are prepared by acylation of the amino groups of 5,10,15,20-tetra(p-aminophenyl)porphyrin by 9-carboranyl acetyl chloride. From the analysis of the absorption and fluorescence spectra, it is concluded that covalent linking of carborane molecules to the tetraphenylporphyrin molecule significantly changes the self-conjugated pi-system of the porphyrin macrocycle: positions of maxima of absorption and fluorescence spectra shift to the red region by 3-8 nm; the halfwidths of these bands are broadened by 2.5-5.0 nm; the relative intensity of the bands I-IV also changes. The fluorescence decay kinetics of the carborane derivatives are biexponential. According to the experimental data and model simulation, it is concluded that the intramolecular electron transfer proceeds from the porphyrin excited part of the molecule to carboranyls with a rate constant of 415 ps(-1) and efficiency of 0.16-0.8. Recombination of separated charges occurs within 1.4 ns.

The influence of structural-dynamic organization of RC from purple bacterium rhodobacter sphaeroides on picosecond stages of photoinduced reactions

Effects of the hydrogen bond network on the rate constants of energy migration (km), charge separation (ke), electron transfer to QA (kQ) and P+I- recombination in RC of Rhodobacter sphaeroides were analysed in control and modified RC preparations at different temperatures. Modification of RC were made by the addition of 40% v/v DMSO. The rate constants km, ke, kQ were evaluated from pump-and-probe measurements of the absorption difference kinetics at 665 nm corresponding to BPhL- formation and subsequent electron transfer to QA. For the investigation of P+I- recombination a primary quinone acceptor was pre-reduced in the dark by adding of 1 mg/ml of dithionite and 1 mM sodium ascorbate. Recombination kinetics were measured at 665 and 870 nm. The numerical analysis of the temperature dependence of ke and kQ was performed on the basis of the model proposed by Kakitani and Kakitani (T. Kakitani and H. Kakitani (1981), Biochim. Biophys. Acta, 635, 498-514). It was found that: (a) in control samples the molecular rate constants km, ke and kQ were about (3.4 ps)-1, (4.5 ps)-1 and (200 ps)-1, respectively; (b) under modification by DMSO these rates decrease up to (5.3 ps)-1, (10.3 ps)-1 and (500 ps)-1, respectively; (c) as the temperature drops from 300 K to 77 K the rate constant km decreases by 1.8 times in control and by 3.2 times in modified samples. In contrast to the observed km changes the increase in ke and kQ values by 2 and more times under cooling was found in control and modified RC; (d) in control preparations with QA acceptor pre-reduced in the dark the lowering of the temperature caused the increase in the time of P+I- recombination from 10 to 20 ns. After DMSO modification the kinetics of charge recombination in RC was biexponential at room temperature with tau=10 ns and tau1=0.8 ns, and at 77 K with tau=20 ns and tau1=0.6 ns, correspondingly. The results obtained reveal that in RC isolated from Rb. sphaeroides the processes of energy migration, charge separation, electron transfer to QA and ion-radical pair P+I- recombination depend on the state of hydrogen bonds of water-protein structure. Fast relaxation processes in RC structure including polarization of H-containing molecules in the surrounding of electron carriers can accept electron energy dissipated at the initial steps of energy and electron transfer. Copyright 1998 Elsevier Science B.V. All rights reserved.

Energetics and mechanisms of high efficiency of charge separation and electron transfer processes in Rhodobacter sphaeroides reaction centers

Effects of environmental changes due to D(2)O/H(2)O substitution and cryosolvent addition on the energetics of the special pair and the rate constants of forward and back electron transfer reactions in the picosecond-nanosecond time domain have been studied in isolated reaction centers (RC) of the anaxogenic purple bacterium Rhodobacter sphaeroides. The following results were obtained: (i). replacement of H(2)O by D(2)O or addition of either 70% (v/v) glycerol or 35% (v/v) DMSO do not influence the absorption spectra; (ii). in marked contrast to this invariance of absorption, the maxima of fluorescence spectra are red shifted relative to control by 3.5, 6.8 and 14.5 nm for RCs suspended in glycerol, D(2)O or DMSO, respectively; (iii). D(2)O/H(2)O substitution or DMSO addition give rise to an increase of the time constants of charge separation (tau(e)), and Q(A)(-) formation (tau(Q)) by a factors of 2.5-3.1 and 1.7-2.5, respectively; (iv). addition of 70% glycerol is virtually without effect on the values of tau(e) and tau(Q); (v). the midpoint potential E(m) of P/P(+) is shifted by about 30 and 45 mV towards higher values by addition of 70% glycerol and 35% DMSO, respectively, but remains invariant to D(2)O/H(2)O exchange; and (vi). an additional fast component with tau(1)=0.5-0.8 ns in the kinetics of charge recombination P(+)H(A)(-)-->P*(P)H(A) emerges in RC suspensions modified either by D(2)O/H(2)O substitution or by DMSO treatment. The results have been analysed with special emphasis on the role of deformations of hydrogen bonds for the solvation mechanism of nonequilibrium states of cofactors. Reorientation of hydrogen bonds provides the major contribution of the very fast environmental response to excitation of the special pair P. The Gibbs standard free energy gap between 1P* and P(+)B(A)(-) due to solvation is estimated to be approximately 70, 59 and 48 meV for control, D(2)O- and DMSO-treated RC samples, respectively.

Purple-bacterial photosynthetic reaction centers and quantum‐dot hybrid‐assemblies in lecithin liposomes and thin films

Quantum dots (QDs) absorb ultraviolet and long-wavelength visible light energy much more efficiently than natural bacterial light-harvesting proteins and can transfer the excitation energy to photosynthetic reaction centers (RCs). Inclusion of RCs combined with QDs as antennae into liposomes opens new opportunities for using such hybrid systems as a basis for artificial energy-transforming devices that potentially can operate with greater efficiency and stability than devices based only on biological components or inorganic components alone. RCs from Rhodobacter sphaeroides and QDs (CdSe/ZnS with hydrophilic covering) were embedded in lecithin liposomes by extrusion of a solution of multilayer lipid vesicles through a polycarbonate membrane or by dialysis of lipids and proteins dispersed with excess detergent. The efficiency of RC and QD interaction within the liposomes was estimated using fluorescence excitation spectra of the photoactive bacteriochlorophyll of the RCs and by measuring the fluorescence decay kinetics of the QDs. The functional activity of the RCs in hybrid complexes was fully maintained, and their stability was even increased. The efficiency of energy transfer between QDs and RCs and conditions of long-term stability of function of such hybrid complexes in film preparations were investigated as well. It was found that dry films containing RCs and QDs, maintained at atmospheric humidity, are capable of maintaining their functional activity for at least some months as judged by measurements of their spectral characteristics, efficiency of energy transfer from QDs to RCs and RC electron transport activity. Addition of trehalose to the films increases the stability further, especially for films maintained at low humidity. These stable hybrid film structures are promising for further studies towards developing new phototransformation devices for biotechnological applications.

The rate of Qx→Qy relaxation in bacteriochlorophylls of reaction centers from Rhodobacter sphaeroides determined by kinetics of the ultrafast carotenoid bandshift

Transient absorption changes induced by excitation of isolated reaction centers (RCs) from Rhodobacter sphaeroides with 600nm laser pulses of 20fs (full width at half maximum) were monitored in the wavelength region of 420-560nm. The spectral features of the spectrum obtained are characteristic for an electrochromic band shift of the single carotenoid (Car) molecule spheroidene, which is an integral constituent of these RCs. This effect is assigned to an electrochromic bandshift of Car due to the local electric field of the dipole moment formed by electronic excitation of bacteriochlorophyll (BChl) molecule(s) in the neighborhood of Car. Based on the known distances between the pigments, the monomeric BChl (B(B)) in the inactive B-branch is inferred to dominate this effect. The excitation of B(B) at 600nm leads to a transition into the S(2) state (Q(x) band), which is followed by rapid internal conversion to the S(1) state (Q(y) band), thus leading to a change of strength and orientation of the dipole moment, i.e., of the electric field acting on the Car molecule. Therefore, the time course of the electrochromic bandshift reflects the rate of the internal conversion from S(2) to S(1) of B(B). The evaluation of the kinetics leads to a value of 30fs for this relaxation process. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: from Natural to Artificial.

The nature of oscillations in the kinetics of electron transfer in the reaction center of purple bacteria.

87 The reaction centers (RCs) of purple bacteria are natural nanostructures able to transform electron excitation energy into the energy of separated charges with a high efficiency (~100%). The RC main unit is formed of two protein subunits, L and M, with four bacteriochlorophyll (BCl) molecules, two bacteriopheophytin ( H A and H B ) molecules, and two quinone ( Q A and Q B ) molecules attached to them. An iron atom is localized between the quinones. In turn, two of the four BCl molecules form a special pair, the primary electron donor P. Spatial organization of the Rhodobacter sphaeroides RC has been determined with a resolution of 2.65 A [1]. Study of the interaction between the excited states and the charge transfer states of RC cofactors provided for discovering oscillations in the kinetics of stimulated luminescence in the R. sphaeroides RC at the excitation at Q y absorption band of the special pair [2, 3]. Shuvalov et al. studied the oscillations in the absorption band of the reduced intermediate acceptor [4, 5]. These oscillations were explained by a wave packet formed on the potential energy surface for the interaction between the special pair and BCl in the active photosynthesis chain during electron transfer. The oscillation data were described by the Redfield super

Temperature Dependence of Tryptophan Fluorescence Lifetime in Aqueous Glycerol and Trehalose Solutions

The temperature dependences of tryptophan fluorescence decay kinetics in aqueous glycerol and 1 M trehalose solutions were examined. The fluorescence decay kinetics were recorded in the spectral region of 292.5–417.5 nm with nanosecond time resolution. The kinetics curves were approximated by the sum of three exponential terms, and the spectral distribution (DAS) of these components was determined. An antisymbatic course of fluorescence decay times of two (fast and medium) components in the temperature range from –60 to +10°C was observed. The third (slow) component showed only slight temperature dependence. The antisymbatic behavior of fluorescence lifetimes of the fast and medium components was explained on the assumption that some of the excited tryptophan molecules are transferred from a short-wave-length B-form with short fluorescence lifetime to a long-wavelength R-form with an intermediate fluorescence lifetime. This transfer occurred in the indicated temperature range.

A comparison of the temperature dependence of charge recombination in the ion-radical pair P870+QA - and tryptophan fluorescence in the photosynthetic reaction centers of Rhodobacter sphaeroides

The temperature dependences of the charge-recombination rate in the ion-radical pair P870+QA- in photosynthetic reaction centers of Rhodobacter sphaeroides were investigated. Recombination kinetics were measured in the individual absorption bands of the donor (600 nm) and an electron acceptor (335 and 420–450 nm) for the reaction center in the water–glycerol and trehalose environment after freezing preparations to–180°С in the dark and on the actinic light and after their subsequent heating. In similar conditions the fluorescence lifetime of tryptophanyls in reaction centers (λreg = 325 and 345 nm), which is an internal indicator of the dynamic state of the protein matrix, was measured. A correlation between the temperature dependences of functional and dynamic parameters of reaction centers in different solvents was shown. The differences in the average fluorescence lifetime of tryptophanyls in reaction centers of preparations frozen in the dark or on the actinic light were found. These results are explained due to transitions of reaction centers between different conformational states and processes of proton relaxation in the structure of the hydrogen bonds in the environment of reaction-center cofactors.

Covalently linked hybrid structures of semiconductor nanocrystals and allophycocyanin

Covalently linked hybrid structures of semiconductor nanocrystals (CdSe/ZnS quantum dots) as an inorganic component with a fluorescence maximum at 620 nm and photosynthetic protein allophycocyanin (APC) as an organic part are created. It is found out that CdSe/ZnS quantum dots form stable complexes with APC through covalent bonding in aqueous solutions. It is shown that the efficiency of electronic excitation energy transfer (EET) in such systems may be significantly enhanced under the conditions at which the monomerization of allophycocyanin trimers occurs. In this paper the EET efficiency is evaluated under differing experimental conditions (pH, temperature, and presence of NaSCN) for hybrid systems obtained by the self-assembling of components via electrostatic interactions, as well as via covalent linking. Under the most optimal conditions, there is a 20-fold increase in the APC fluorescence after the excitation of QDs due to the more efficient EET for the covalently linked components compared to the complexes obtained by the self-assemblage. The obtained covalently linked hybrid structures unfold new opportunities for their practical use as fluorescent markers, hybrid photosensors, and structural elements in photovoltaic devices.

Generation of radical form of dipyridamole at illumination of photosynthetic reaction centers of Rb. sphaeroides

The study of the effect of vasodilator, antiplatelet agent, and inhibitor P-glycoprotein dipyridamole (DIP) on the functioning of the transmembrane protein of the reaction center (RC) of Rb. sphaeroides showed that the activation of RC by constant light generates the DIP radical cation, which significantly affects the kinetics of recombination of charges divided between photoactive bacteriochlorophyll and quinone acceptors. Thus, the antioxidant properties of DIP may affect the functional activity of membrane proteins, and this apparently should be taken into account in the studies of the mechanisms of therapeutic action of this drug.