Bacterial Photosynthetic Reaction Centers Research Articles

Magnetic field effect on singlet oxygen production in a biochemical system

The yield of singlet oxygen sensitized by chemically modified, carotenoidless bacterial photosynthetic reaction centres and the ensuing oxidative damage are both shown to be magnetic field-dependent.

Control of Chemical Equilibrium by Noise

We formulate a theory of reactive transport in the presence of environmental fluctuations that restart a transfer attempt, such as, for instance, during charge transfer through an intermediate state on a fluctuating bridge. It is shown that the rate and statistics of the fluctuations affect the steady-state populations of the reactants and products and the rate at which the steady state is approached. We give explicit examples in which the reactant/product populations can be inverted by varying the rate of environmental fluctuations. We thus propose a design principle of a controllable molecular scale device. We speculate that when noise-controlled charge transfer takes place in nature, as during electron transport in bacterial photosynthetic reaction centers, such a design can be used to maintain functionality over a wide temperature range and would be favored by evolution.

Theory of Optical Spectra Involving Charge Transfer States: Dynamic Localization Predicts a Temperature Dependent Optical Band Shift

The influence of charge transfer states on the optical line shape of chromophore complexes is investigated in a minimal model that includes a coupling between an excited state and an optically dark charge transfer state. In the calculations of the absorption spectrum, an intensity borrowing by the charge transfer state, strong vibrational sidebands, and a temperature dependent shift of optical transitions are obtained. The theory is applied to the bacterial photosynthetic reaction center to explain a 30 nm blueshift of the low-energy absorption band with increasing temperature.

Protein/lipid interaction in the bacterial photosynthetic reaction center: phosphatidylcholine and phosphatidylglycerol modify the free energy levels of the quinones.

The role of characteristic phospholipids of native membranes, phosphatidylcholine (PC), phosphatidylglycerol (PG), and cardiolipin (CL), was studied in the energetics of the acceptor quinone side in photosynthetic reaction centers of Rhodobacter sphaeroides. The rates of the first, k(AB)(1), and the second, k(AB)(2), electron transfer and that of the charge recombination, k(BP), the free energy levels of Q(A)(-)Q(B) and Q(A)Q(B)(-) states, and the changes of charge compensating protein relaxation were determined in RCs incorporated into artificial lipid bilayer membranes. In RCs embedded in the PC vesicle, k(AB)(1) and k(AB)(2) increased (from 3100 to 4100 s(-1) and from 740 to 3300 s(-1), respectively) and k(BP) decreased (from 0.77 to 0.39 s(-1)) compared to those measured in detergent at pH 7. In PG, k(AB)(1) and k(BP) decreased (to values of 710 and 0.26 s(-1), respectively), while k(AB)(2) increased to 1506 s(-1) at pH 7. The free energy between the Q(A)(-)Q(B) and Q(A)Q(B)(-) states decreased in PC and PG (DeltaG degrees (Q)A-(Q)B(-->)(Q)A(Q)B- = -76.9 and -88.5 meV, respectively) compared to that measured in detergent (-61.8 meV). The changes of the Q(A)/Q(A)(-) redox potential measured by delayed luminescence showed (1) a differential effect of lipids whether RC incorporated in micelles or vesicles, (2) an altered binding interaction between anionic lipids and RC, (3) a direct influence of PC and PG on the free energy levels of the primary and secondary quinones probably through the intraprotein hydrogen-bonding network, and (4) a larger increase of the Q(A)/Q(A)(-) free energy in PG than in PC both in detergent micelles and in single-component vesicles. On the basis of recent structural data, implications of the binding properties of phospholipids to RC and possible interactions between lipids and electron transfer components will be discussed.

Light-driven conformational changes in a bacterial photosynthetic reaction centre

Light-driven conformational changes in a bacterial photosynthetic reaction centre

Long-lived charge-separated state generated in a ferrocene-meso,meso-linked porphyrin trimer-fullerene pentad with a high quantum yield.

A meso,meso-linked porphyrin trimer, (ZnP)3, as a light-harvesting chromophore, has been incorporated for the first time into a photosynthetic multistep electron-transfer model including ferrocene (Fc) as an electron donor and fullerene (C60) as an electron acceptor, to construct the ferrocene-meso,meso-linked porphyrin trimer-fullerene system Fc-(ZnP)3-C60. Photoirradiation of Fc-(ZnP)3-C60 results in photoinduced electron transfer from both the singlet and triplet excited states of the porphyrin trimer, 1(ZnP)3* and 3(ZnP)3*, to the C60 moiety to produce the porphyrin trimer radical cation-C60 radical anion pair, Fc-(ZnP)3*+-C60*-. Subsequent formation of the final charge-separated state Fc+-(ZnP)3-C60*- was confirmed by the transient absorption spectra observed by pico- and nanosecond time-resolved laser flash photolysis. The final charge-separated state decays, obeying first-order kinetics, with a long lifetime (0.53 s in DMF at 163 K) that is comparable with that of the natural bacterial photosynthetic reaction center. More importantly, the quantum yield of formation of the final charge-separated state (0.83 in benzonitrile) remains high, despite the large separation distance between the Fc+ and C60*- moieties. Such a high quantum yield results from efficient charge separation through the porphyrin trimer, whereas a slow charge recombination is associated with the localized porphyrin radical cation in the porphyrin trimer. The light-harvesting efficiency in the visible region has also been much improved in Fc-(ZnP)3-C60 because of exciton coupling in the porphyrin trimer as well as an increase in the number of porphyrins.

Pulsed EPR/ENDOR Characterization of the Cu2+ Surface Site in Photosynthetic Bacterial Reaction Centers

X-band pulsed electron paramagnetic resonance was used to elucidate details of the Cu2+ surface binding site involved in QA-QB → QAQB- electron transfer in photosynthetic bacterial reaction centers (RCs). Electron spin−echo envelope modulation (ESEEM) spectroscopy was employed to measure the hyperfine and electric quadrupole interactions of weakly coupled 14N nuclei, and X-band pulsed electron nuclear double resonance (ENDOR) spectroscopy was used to probe strongly coupled 14N nuclei. The 14N ESEEM observed for Cu2+ at the surface site of RCs results from a single histidine ligand. In addition, the ENDOR spectra reveal the presence of one more nitrogen ligand in the Cu2+ coordination sphere that we tentatively assign as a peptide backbone nitrogen. The similarity of the ESEEM and ENDOR spectra for RCs from Rb. sphaeroides, Rb. capsulatus, and Blc. viridis suggests a universal intrinsic structural site. These results indicate that Cu2+ has a different coordination environment from the crystallographically ...

Spin distribution and the location of protons in paramagnetic proteins.

Two current frontiers in EPR research are high-field ( nu0 > 70 GHz, B0 > 2.5 T ) electron paramagnetic resonance (EPR) and high-field electron-nuclear double resonance (ENDOR). This review focuses on recent advances in high-field ENDOR and its applications to the study of proteins containing native paramagnetic sites. It concentrates on two aspects; the first concerns the determination of the location of protons and is related to the site geometry, and the second focuses on the spin density distribution within the site, which is inherent to the electronic structure. Both spin density and proton locations can be derived from ligand hyperfine couplings determined by ENDOR measurements. A brief description of the experimental methods is presented along with a discussion of the advantages and disadvantages of high-field ENDOR compared with conventional X-band (~ 9.5 GHz) experiments. Specific examples of both protein single crystals and frozen solutions are then presented. These include the determination of the coordinates of water ligand protons in the Mn(II) site of concanavalin A, the detection of hydrogen bonds in a quinone radical in the bacterial photosynthetic reaction center as well as in the tyrosyl radical in ribonuclease reductase, and the study of the spin distribution in copper proteins. The copper proteins discussed are the type I copper of azurin and the binuclear CuA center in a number of proteins. The last part of the review presents a brief discussion of the interpretation of hyperfine couplings using quantum chemical calculations, primarily density functional theory (DFT) methods. Such methods are becoming an integral part of the data analysis tools, as they can facilitate signal assignment and provide the ultimate relation between the experimental hyperfine couplings and the electronic wave function.

Variation of Ser-L223 hydrogen bonding with the QB redox state in reaction centers from Rhodobacter sphaeroides.

Ser-L223 is close to ubiquinone (Q(B)) in the B-branch of the bacterial photosynthetic reaction center (bRC) from Rhodobacter (Rb) sphaeroides. Therefore, the presence of a hydrogen bond (H bond) between the two was naturally proposed from the crystal structure. The hydrogen bonding pattern of Q(B) from the light-exposed structure was studied by generating hydrogen atom coordinates based on the CHARMM force field. In the Q(B) neutral charge state (Q(B)(0)), no H bond was found between the oxygen of the OH group from Ser-L223 and the carbonyl oxygen of Q(B) that is distal to the non-heme iron. In the reduced state (Q(B)(-)), however, Ser-L213 was found to form an H bond with Q(B) only when Asp-L213 is protonated by more than 0.75 H(+). This indicates the significance of the protonation of Asp-L213 in forming an H bond between Ser-L223 and Q(B). We found that the driving force to form the H bond between Ser-L223 and Q(B) is enhanced by the positively charged Arg-L217. The calculated Q(B) redox potentials with or without this H bond discriminated two ET rates, which are close to the faster and slower time phases observed in UV-Vis and FTIR studies. Together with the calculated redox potential of the quinones, this H-bond formation could play a key role in conformational gating for the ET process from Q(A) to Q(B).

Dependence of Photosynthetic Electron-Transfer Kinetics on Temperature and Energy in a Density-Matrix Model

We use a multidimensional density-matrix model to explore the temperature dependence of electron transfer from the excited singlet state of the primary electron donor (P*) to the neighboring bacteriochlorophyll (B A ) in photosynthetic bacterial reaction centers. This reaction, which has the unusual property of increasing in rate with decreasing temperature, occurs too rapidly to be treated reliably by approaches that assume thermal equilibration of the vibrational levels of the reactant electronic state. In the density-matrix treatment, the frequencies and displacements of the vibrational modes that are coupled to electron transfer, and the microscopic time constants for transitions between different vibrational states, are obtained from molecular-dynamics simulations by the dispersed-polaron (spin-boson) approach. The electron-transfer dynamics are simulated by integrating the stochastic Liouville equation following excitation of the system with a short pulse of light. In this model, the increase in the rate with decreasing temperature depends strongly on the fact that electron transfer occurs more rapidly than vibrational thermalization. The high rate of electron transfer also affects the dependence of the kinetics on the energy difference between the reactant and product electronic states (ΔE°), making the optimal value of -AE° smaller than the reorganization energy. This interesting effect can be rationalized qualitatively in simple semiclassical and quantum mechanical models.

Electrochromic Shift of Chlorophyll Absorption in Photosystem I from Synechocystis sp. PCC 6803: A Probe of Optical and Dielectric Properties around the Secondary Electron Acceptor

Nanosecond absorption dynamics at ∼685 nm after excitation of photosystem I (PS I) from Synechocystis sp. PCC 6803 is consistent with electrochromic shift of absorption bands of the Chl a pigments in the vicinity of the secondary electron acceptor A 1. Based on experimental optical data and structure-based simulations, the effective local dielectric constant has been estimated to be between 3 and 20, which suggests that electron transfer in PS I is accompanied by considerable protein relaxation. Similar effective dielectric constant values have been previously observed for the bacterial photosynthetic reaction center and indicate that protein reorganization leading to effective charge screening may be a necessary structural property of proteins that facilitate the charge transfer function. The data presented here also argue against attributing redmost absorption in PS I to closely spaced antenna chlorophylls (Chls) A38 and A39, and suggest that optical transitions of these Chls, along with that of connecting chlorophyll (A40) lie in the range 680–695 nm.

X-Ray Structure Determination of Three Mutants of the Bacterial Photosynthetic Reaction Centers from Rb. sphaeroides: Altered Proton Transfer Pathways

In the photosynthetic reaction center (RC) from Rhodobacter sphaeroides, the reduction of a bound quinone molecule Q B is coupled with proton uptake. When Asp-L213 is replaced by Asn, proton transfer is inhibited. Proton transfer was restored by two second-site revertant mutations, Arg-M233→Cys and Arg-H177→His. Kinetic effects of Cd 2+ on proton transfer showed that the entry point in revertant RCs to be the same as in the native RC. The structures of the parental and two revertant RCs were determined at resolutions of 2.10, 1.80, and 2.75 Å. From the structures, we were able to delineate alternate proton transfer pathways in the revertants. The main changes occur near Glu-H173, which allow it to substitute for the missing Asp-L213. The electrostatic changes near Glu-H173 cause it to be a good proton donor and acceptor, and the structural changes create a cavity which accommodates water molecules that connect Glu-H173 to other proton transfer components.

Potential Energy Landscape for Conformationally Gated Secondary Ubiquinone Binding in the Photosynthetic Reaction Center of Rhodobacter Sphaeroides

Migration of the secondary ubiquinone, UQB, to a site proximal to His-L190 was proposed to be rate-determining for electron transfer in bacterial photosynthetic reaction centers (shown). This study shows that 1) the proximal binding site is the global minimum on the potential energy landscape, 2) multiple minima exist, in qualitative agreement with various binding sites observed in X-ray diffraction studies, and 3) the energy barrier for direct migration of UQB and migration gated by a protein conformational change both agree with the activation energy measured for electron transfer.

Electron Transfer between Quinones in Photosynthetic Reaction Centers

The mechanism of electron transfer (ET) from the primary to the secondary quinone of bacterial photosynthetic reaction centers is discussed on the basis of theoretical computations of the minimum energy nuclear configurations and ET coupling elements, and quantum dynamic simulations of elementary reaction steps. For ET to occur via tunneling, unreasonably high values of the electronic coupling elements or very stringent energy conditions, i.e., tight degeneracy (within a few cm-1) between the initial and final vibronic states, are necessary, both for the direct and through-bridge (superexchange) routes. The assumption of tight degeneracy significantly slows down the process so that other competitive processes, such as proton transfer from the H-bonded HisM219 to the primary quinone, can take place. All these results suggest that the iron−histidine bridge can play an important role in the ET mechanism.

A Molecular Tetrad Allowing Efficient Energy Storage for 1.6 s at 163 K

In a novel molecular ferrocene−zinc porphyrin−zinc porphyrin−fullerene (Fc−ZnP−ZnP−C60) tetrad, the longest lifetime of a charge-separated state ever reported in an artificial photosynthetic reaction center (1.6 s in DMF at 163 K) has been attained. This lifetime is comparable, for example, to the lifetime (∼1 s) of the bacteriochlorophyll dimer radical cation ((Bchl)2•+)−secondary quinone radical anion (QB•-) ion pair in the bacterial photosynthetic reaction centers. The present far distant radical ion pair is formed with a quantum yield of 34%. The radical ion pair Fc+−ZnP−ZnP−C60•- produced by photoinduced electron transfer was detected by means of transient absorption spectra as well as ESR spectra. Both the lifetime and the quantum yield of the final charge-separated state are improved in Fc−ZnP−ZnP−C60 relative to the corresponding ferrocene−zinc porphyrin−free base porphyrin−fullerene (Fc−ZnP−H2P−C60) tetrad.

2P248 光合成バクテリア反応中心のBacteriopheophytinからprimary quinoneへの電子移動における非弾性トンネル現象(光生物 B) 光合成)

2P248 光合成バクテリア反応中心のBacteriopheophytinからprimary quinoneへの電子移動における非弾性トンネル現象(光生物 B) 光合成)

Spontaneous emulsification of detergent solubilized reaction center: protein conformational changes precede droplet growth

We show that ionization of a pH-sensitive detergent, DDAO, bound to a bacterial photosynthetic reaction center (RC), induces reversible emulsification of the protein over a narrow acidic pH range, resulting in stable micrometric RC-surfactant droplets. Electrostatic interactions play a key role in the phase separation process, as shown by a systematic analysis of ionic strength effects and by the use of a cationic detergent (DTAB) that mimics, also at basic pH, the ionized form of DDAO. Under all the conditions we tested, phase segregation seems to be coupled to a 15 nm blue-shift of the low energy absorption band of the primary electron donor P of the RC. This spectral change strongly suggests that surfactant-protein interactions leading to phase separation also induce a conformational transition of the RC. Time-resolved visible-NIR spectra recorded during the emulsification process reveal that the conformational change probed by P spectral shift is always faster than droplets formation. In line with these observations, phase segregation affects charge recombination kinetics following RC photoexcitation, as well as electron transfer from soluble cytochrome c2 to the photoxidized primary donor P+.

A density-matrix model of photosynthetic electron transfer with microscopically estimated vibrational relaxation times

Abstract The dispersed-polaron (spin-boson) model is reviewed briefly and then used to develop a density-matrix model for studies of electron transfer in condensed phases. The frequencies and Franck–Condon factors for solvent vibrational modes that are coupled to electron transfer are obtained from molecular dynamics (MD) simulations by the dispersed-polaron treatment. Microscopic rate constants for vibrational relaxations, dephasing and coherence transfer between the solvent modes are obtained by fitting the time dependence of the solvent coordinates in the density-matrix treatment to the corresponding time dependence obtained from molecular-dynamics simulations with a classical linear-response approximation. This is done by adjusting a single parameter, the time constant for thermal equilibration of the two lowest levels of a solvent mode (T10). The model thus focuses on the coupling between solvent modes, rather than on the more widely studied coupling of solute modes by the thermal bath. The resulting density-matrix model is used to examine vibronic coupling in the initial electron-transfer step in photosynthetic bacterial reaction centers. Values of T10 in the range of 1–2 ps are consistent with molecular-dynamics simulations of the time-dependent energy gap between the reactant and product states (P* and P+B−), and also with the damping of coherent vibrational motions that are seen experimentally after excitation of reaction centers with a short pulse of light. In both the density-matrix model and the MD simulations, the autocorrelation function of the energy gap also has a decay component with a time constant of about 50 fs, which we ascribe to the group dephasing of oscillatory motions at many different frequencies. This component is insensitive to vibrational relaxations and is largely irrelevant to the electron-transfer dynamics. Using values of T10 in the range of 1–2 ps, a model with five vibrational modes reproduces the main features of electron transfer from P* to B, including stepwise formation of the product during the period when the system retains vibrational coherences. Although the rate does not depend strongly on whether P* is prepared coherently or incoherently, speeding up vibrational relaxations decreases the rate. At least part of the adverse effect of rapid relaxations can be viewed as a manifestation of the quantum Zeno paradox, which arises when off-diagonal elements of the density matrix decay very rapidly.

Intervalence Band Stark Effect of the Special Pair Radical Cation in Bacterial Photosynthetic Reaction Centers

The Stark spectrum of the intervalence band of the special pair radical cation in bacterial photosynthetic reaction centers is presented. This spectrum, centered at 2600 cm-1, is modeled using a general treatment of intervalence band Stark effects based on a two-state vibronic coupling model of mixed-valency. The observed line shape can be predicted using values for the electronic coupling, driving force, and reorganization energy along a single antisymmetric mode as derived from previous measurements. The model can be used to estimate the difference between the electric dipole moments of the two hole-localized electronic states of a mixed-valence complex, ΔμCT, even if the dipole strength of its IVB is unknown or affected by intensity borrowing by other electronic transitions. We determine the value of ΔμCT to be 33 ± 5 D/f for the special pair radical cation in wild-type Rhodobacter sphaeroides reaction centers, where f is the scalar approximation to the local field correction tensor. This value of ΔμCT is less than would be predicted from the separation between the magnesium ions of the special pair, yet large enough to suggest that the observed dipole strength of its intervalence band is strongly affected by intensity borrowing.

Charge Recombination and Protein Dynamics in Bacterial Photosynthetic Reaction Centers Entrapped in a Sol-Gel Matrix

Many proteins can be immobilized in silica hydrogel matrices without compromising their function, making this a suitable technique for biosensor applications. Immobilization will in general affect protein structure and dynamics. To study these effects, we have measured the P+QA− charge recombination kinetics after laser excitation of QB-depleted wild-type photosynthetic reaction centers from Rhodobacter sphaeroides in a tetramethoxysilane (TMOS) sol-gel matrix and, for comparison, also in cryosolvent. The nonexponential electron transfer kinetics observed between 10 and 300K were analyzed quantitatively using the spin boson model for the intrinsic temperature dependence of the electron transfer and an adiabatic change of the energy gap and electronic coupling caused by protein motions in response to the altered charge distributions. The analysis reveals similarities and differences in the TMOS-matrix and bulk-solvent samples. In both preparations, electron transfer is coupled to the same spectrum of low frequency phonons. As in bulk solvent, charge-solvating protein motions are present in the TMOS matrix. Large-scale conformational changes are arrested in the hydrogel, as evident from the nonexponential kinetics even at room temperature. The altered dynamics is likely responsible for the observed changes in the electronic coupling matrix element.