BChl Dimer Research Articles

Probing ligands to reaction centers to limit the photocycle in photosynthetic bacterium Rubrivivax gelatinosus

Light-induced electron flow between reaction center and cytochrome bc1 complexes is mediated by quinones and electron donors in purple photosynthetic bacteria. Upon high-intensity excitation, the contribution of the cytochrome bc1 complex is limited kinetically and the electron supply should be provided by the pool of reduced electron donors. The kinetic limitation of electron shuttle between reaction center and cytochrome bc1 complex and its consequences on the photocycle were studied by tracking the redox changes of the primary electron donor (BChl dimer) via absorption change and the opening of the closed reaction center via relaxation of the bacteriochlorophyll fluorescence in intact cells of wild type and pufC mutant strains of Rubrivivax gelatinosus. The results were simulated by a minimum model of reversible binding of different ligands (internal and external electron donors and inhibitors) to donor and acceptor sides of the reaction center. The calculated binding and kinetic parameters revealed that control of the rate of the photocycle is primarily due to 1) the light intensity, 2) the size and redox state of the donor pool, and 3) the unbinding rates of the oxidized donor and inhibitor from the reaction center. The similar kinetics of strains WT and pufC lacking the tetraheme cytochrome subunit attached to the reaction center raise the issue of the physiological importance of this subunit discussed from different points of view. SignificanceA crucial factor for the efficacy of electron donors in photosynthetic photocycle is not just the substantial size of the pool and large binding affinity (small dissociation constant KD = koff/kon) to the RC, but also the mean residence time (koff)−1 in the binding pocket. This is an important parameter that regulates the time of re-activation of the RC during multiple turnovers. The determination of koff has proven challenging and was performed by simulation of widespread experimental data on the kinetics of P+ and relaxation of fluorescence. This work is a step towards better understanding the complex pathways of electron transfer in proteins and simulation-based design of more effective electron transfer components in natural and artificial systems.

X-ray structure of the Rhodobacter sphaeroides reaction center with an M197 Phe→His substitution clarifies the properties of the mutant complex.

The first steps of the global process of photosynthesis take place in specialized membrane pigment-protein complexes called photosynthetic reaction centers (RCs). The RC of the photosynthetic purple bacterium Rhodobacter sphaeroides, a relatively simple analog of the more complexly organized photosystem II in plants, algae and cyanobacteria, serves as a convenient model for studying pigment-protein interactions that affect photochemical processes. In bacterial RCs the bacteriochlorophyll (BChl) dimer P serves as the primary electron donor, and its redox potential is a critical factor in the efficient functioning of the RC. It has previously been shown that the replacement of Phe M197 by His strongly affects the oxidation potential of P (E m P/P+), increasing its value by 125 mV, as well as increasing the thermal stability of RC and its stability in response to external pressure. The crystal structures of F(M197)H RC at high resolution obtained using various techniques presented in this report clarify the optical and electrochemical properties of the primary electron donor and the increased resistance of the mutant complex to denaturation. The electron-density maps are consistent with the donation of a hydrogen bond from the imidazole group of His M197 to the C2-acetyl carbonyl group of BChl PB. The formation of this hydrogen bond leads to a considerable out-of-plane rotation of the acetyl carbonyl group and results in a 1.2 Å shift of the O atom of this group relative to the wild-type structure. Besides, the distance between BChl PA and PB in the area of pyrrole ring I was found to be increased by up to 0.17 Å. These structural changes are discussed in association with the spectral properties of BChl dimer P. The electron-density maps strongly suggest that the imidazole group of His M197 accepts another hydrogen bond from the nearest water molecule, which in turn appears to form two more hydrogen bonds to Asn M195 and Asp L155. As a result of the F(M197)H mutation, BChl PB finds itself connected to the extensive hydrogen-bonding network that pre-existed in wild-type RC. Dissimilarities in the two hydrogen-bonding networks near the M197 and L168 sites may account for the different changes of the E m P/P+ in F(M197)H and H(L168)F RCs. The involvement of His M197 in the hydrogen-bonding network also appears to be related to stabilization of the F(M197)H RC structure. Analysis of the experimental data presented here and of the data available in the literature points to the fact that the hydrogen-bonding networks in the vicinity of BChl dimer P may play an important role in fine-tuning the redox properties of the primary electron donor.

The flexible N-terminus of BchL autoinhibits activity through interaction with its [4Fe-4S] cluster and released upon ATP binding

A key step in bacteriochlorophyll biosynthesis is the reduction of protochlorophyllide to chlorophyllide, catalyzed by dark-operative protochlorophyllide oxidoreductase. Dark-operative protochlorophyllide oxidoreductase contains two [4Fe-4S]–containing component proteins (BchL and BchNB) that assemble upon ATP binding to BchL to coordinate electron transfer and protochlorophyllide reduction. But the precise nature of the ATP-induced conformational changes is poorly understood. We present a crystal structure of BchL in the nucleotide-free form where a conserved, flexible region in the N-terminus masks the [4Fe-4S] cluster at the docking interface between BchL and BchNB. Amino acid substitutions in this region produce a hyperactive enzyme complex, suggesting a role for the N-terminus in autoinhibition. Hydrogen–deuterium exchange mass spectrometry shows that ATP binding to BchL produces specific conformational changes leading to release of the flexible N-terminus from the docking interface. The release also promotes changes within the local environment surrounding the [4Fe-4S] cluster and promotes BchL-complex formation with BchNB. A key patch of amino acids, Asp-Phe-Asp (the ‘DFD patch’), situated at the mouth of the BchL ATP-binding pocket promotes intersubunit cross stabilization of the two subunits. A linked BchL dimer with one defective ATP-binding site does not support protochlorophyllide reduction, illustrating nucleotide binding to both subunits as a prerequisite for the intersubunit cross stabilization. The masking of the [4Fe-4S] cluster by the flexible N-terminal region and the associated inhibition of the activity is a novel mechanism of regulation in metalloproteins. Such mechanisms are possibly an adaptation to the anaerobic nature of eubacterial cells with poor tolerance for oxygen.

On the Role of the Special Pair in Photosystems as a Charge Transfer Rectifier.

The special pair, a bacteriochlorophyll a (BChl) dimer found at the core of bacterial reaction centers, is known to play a key role in the functionality of photosystems as a precursor to the photosynthesis process. In this paper, we analyze the inherent affinity of the special pair to rectify the intrapair photo-induced charge transfer (CT). In particular, we show that the molecular environment affects the nuclear geometry, resulting in symmetry breaking between the two possible intrapair CT processes. To this end, we study the relationships of the intrapair CT and the molecular geometry with respect to the effective dielectric constant provided by the molecular environment. We identify the special pair structural feature that breaks the symmetry between the two molecules, leading to CT rectification. Excited state energies, oscillator strengths, and electronic coupling values are obtained via time-dependent density functional theory, employing a recently developed framework based on a screened range-separated hybrid functional within a polarizable continuum model (SRSH-PCM). We analyze the rectification capability of the special pair by calculating the CT rates using a first-principles-based Fermi's golden rule approach.

Unique double concentric ring organization of light harvesting complexes in Gemmatimonas phototrophica.

The majority of life on Earth depends directly or indirectly on the sun as a source of energy. The initial step of photosynthesis is facilitated by light-harvesting complexes, which capture and transfer light energy into the reaction centers (RCs). Here, we analyzed the organization of photosynthetic (PS) complexes in the bacterium G. phototrophica, which so far is the only phototrophic representative of the bacterial phylum Gemmatimonadetes. The isolated complex has a molecular weight of about 800 ± 100 kDa, which is approximately 2 times larger than the core complex of Rhodospirillum rubrum. The complex contains 62.4 ± 4.7 bacteriochlorophyll (BChl) a molecules absorbing in 2 distinct infrared absorption bands with maxima at 816 and 868 nm. Using femtosecond transient absorption spectroscopy, we determined the energy transfer time between these spectral bands as 2 ps. Single particle analyses of the purified complexes showed that they were circular structures with an outer diameter of approximately 18 nm and a thickness of 7 nm. Based on the obtained, we propose that the light-harvesting complexes in G. phototrophica form 2 concentric rings surrounding the type 2 RC. The inner ring (corresponding to the B868 absorption band) is composed of 15 subunits and is analogous to the inner light-harvesting complex 1 (LH1) in purple bacteria. The outer ring is composed of 15 more distant BChl dimers with no or slow energy transfer between them, resulting in the B816 absorption band. This completely unique and elegant organization offers good structural stability, as well as high efficiency of light harvesting. Our results reveal that while the PS apparatus of Gemmatimonadetes was acquired via horizontal gene transfer from purple bacteria, it later evolved along its own pathway, devising a new arrangement of its light harvesting complexes.

Stoichiometry of ATP hydrolysis and chlorophyllide formation of dark-operative protochlorophyllide oxidoreductase from Rhodobacter capsulatus

Dark-operative protochlorophyllide (Pchlide) oxidoreductase (DPOR) is a nitrogenase-like enzyme catalyzing a reduction of the C17 = C18 double bond of Pchlide to form chlorophyllide a (Chlide) in bacteriochlorophyll biosynthesis. DPOR consists of an ATP-dependent reductase component, L-protein (a BchL dimer), and a catalytic component, NB-protein (a BchN–BchB heterotetramer). The L-protein transfers electrons to the NB-protein to reduce Pchlide, which is coupled with ATP hydrolysis. Here we determined the stoichiometry of ATP hydrolysis and the Chlide formation of DPOR. The minimal ratio of ATP to Chlide (ATP/2e–) was 4, which coincides with that of nitrogenase. The ratio increases with increasing molar ratio of L-protein to NB-protein. This profile differs from that of nitrogenase. These results suggest that DPOR has a specific intrinsic property, while retaining the common features shared with nitrogenase.

Different effects of identical symmetry-related mutations near the bacteriochlorophyll dimer in the photosynthetic reaction center of Rhodobacter sphaeroides.

In the bacterial photosynthetic reaction center (RC), asymmetric protein environment of the bacteriochlorophyll (BChl) dimer largely determines the photophysical and photochemical properties of the primary electron donor. Previously, we noticed significant differences in properties of Rhodobacter sphaeroides RCs with identical mutations in symmetry-related positions- I(M206)H and I(L177)H. The substitution I(L177)H resulted in covalent binding of BChl PA with the L-subunit, as well as in 6-coordination of BChl BB, whereas in RC I(M206)H no such changes of pigment-protein interactions were found. In addition, the yield of RC I(M206)H after its isolation from membranes was significantly lower than the yield of RC I(L177)H. This study shows that replacement of amino acid residues in the M203-M206 positions near BChls PB and BA by symmetry-related residues from the L-subunit near BChls PA and BB leads to further decrease in RC amount in the membranes associated obviously with poor assembly of the complex. Introduction of a new hydrogen bond between BChl PB and its protein environment by means of the F(M197)H mutation stabilized the mutant RC but did not affect its low yield. We suggest that the mutation I(M206)H and substitution of amino acid residues in M203-M205 positions could disturb glycolipid binding on the RC surface near BChl BA that is important for stable assembly of the complex in the membrane.

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 6nm 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 RCL(M196)H determined at 2.4Å 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.

Probing the spatial organization of bacteriochlorophyll C by solid-state nuclear magnetic resonance.

Green sulfur bacteria, which live in extremely low-light environments, use chlorosomes to harvest light. A chlorosome is the most efficient, and arguably the simplest, light-harvesting antenna complex, which contains hundreds of thousands of densely packed bacteriochlorophylls (BChls). To harvest light efficiently, BChls in a chlorosome form supramolecular aggregates; thus, it is of great interest to determine the organization of the BChls in a chlorosome. In this study, we conducted a (13)C solid-state nuclear magnetic resonance and Mg K-edge X-ray absorption analysis of chlorosomes from wild-type Chlorobaculum tepidum. The X-ray absorption results indicated that the coordination number of the Mg in the chlorosome must be >4, providing evidence that electrostatic interactions formed between the Mg of a BChl and the carbonyl group or the hydroxyl group of the neighboring BChl molecule. According to the intermolecular distance constraints obtained on the basis of (13)C homonuclear dipolar correlation spectroscopy, we determined that the molecular assembly of BChls is dimer-based and that the hydrogen bonds among the BChls are less extensive than commonly presumed because of the twist in the orientation of the BChl dimers. This paper also reports the first (13)C homonuclear correlation spectrum acquired for carotenoids and lipids-which are minor, but crucial, components of chlorosomes-extracted from wild-type Cba. tepidum.

The site-directed mutation I(L177)H in Rhodobacter sphaeroides reaction center affects coordination of PA and BB bacteriochlorophylls

To explore the influence of the I(L177)H single mutation on the properties of the nearest bacteriochlorophylls (BChls), three reaction centers (RCs) bearing double mutations were constructed in the photosynthetic purple bacterium Rhodobacter sphaeroides, and their properties and pigment content were compared with those of the correspondent single mutant RCs. Each pair of the mutations comprised the amino acid substitution I(L177)H and another mutation altering histidine ligand of BChl PA or BChl BB. Contrary to expectations, the double mutation I(L177)H+H(L173)L does not bring about a heterodimer RC but causes a 46nm blue shift of the long-wavelength P absorbance band. The histidine L177 or a water molecule were suggested as putative ligands for PA in the RC I(L177)H+H(L173)L although this would imply a reorientation of the His backbone and additional rearrangements in the primary donor environment or even a repositioning of the BChl dimer. The crystal structure of the mutant I(L177)H reaction center determined to a resolution of 2.9Å shows changes at the interface region between the BChl PA and the monomeric BChl BB. Spectral and pigment analysis provided evidence for β-coordination of the BChl BB in the double mutant RC I(L177)H+H(M182)L and for its hexacoordination in the mutant reaction center I(L177)H. Computer modeling suggests involvement of two water molecules in the β-coordination of the BChl BB. Possible structural consequences of the L177 mutation affecting the coordination of the two BChls PA and BB are discussed. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: from Natural to Artificial.

Bacteriochlorophyll Excited-State Quenching Pathways in Bacterial Reaction Centers with the Primary Donor Oxidized

One striking feature of bacterial reaction centers is that while they show a high degree of structural symmetry, function is entirely asymmetric: excitation of the primary electron donor, P, a bacteriochlorophyll (BChl) dimer, results almost exclusively in electron transfer along one of the two symmetric electron transfer pathways. Here another functional asymmetry of the reaction center is explored; i.e., the two monomer BChl molecules (B(A) and B(B)) have distinct interactions with P in the oxidized state, P(+). Previous work has suggested that the excited states of both B(A) and B(B) were quenched via energy transfer to P(+) within a few hundred femtoseconds. Here, it is shown that various excitation wavelengths, corresponding to different initial B(A) and B(B) excited states, result in distinct reaction pathways, and which pathway dominates depends both on the initial excited state formed and on the electronic structure of P(+). In particular, it is possible to specifically excite the Q(X) transition of B(B) by using excitation at 495 nm directly into the carotenoid S(2) state which then undergoes energy transfer to B(B). This results in the formation of a new state on the picosecond time scale that is both much longer lived and spectrally different than what one would expect for a simple excited state. Combining results from additional measurements using nonselective 600 or 800 nm excitation of both B(A) and B(B) to the Q(X) or Q(Y) states, respectively, it is found that B(B)* and B(A)* are quenched by P(+) with different kinetics and mechanisms. B(A)* formed using either Q(X) or Q(Y) excitation appears to decay rapidly (∼200 fs) without a detectable intermediate. In contrast, B(B)* formed via Q(X) excitation predominantly generates the long-lived state referred to above via an electron transfer reaction from the Q(X) excited state of B(B) to P(+). This reaction is in competition with intramolecular relaxation of the Q(X) state to the lowest singlet excited state. The Q(Y) excited state of B(B) appears to undergo the electron transfer reaction seen upon Q(X) excitation only to a very limited extent and is largely quenched via energy transfer to P(+). Finally, the ability of P(+) to quench B(B)* depends on the electronic structure of P(+). The asymmetric charge distribution between the two halves of P in the native reaction center is effectively reversed in the mutant HF(L168)/LH(L131), and in this case, the rate of quenching decreases significantly.

Isolation and Characterization of Homodimeric Type-I Reaction Center Complex from Candidatus Chloracidobacterium thermophilum, an Aerobic Chlorophototroph

The recently discovered thermophilic acidobacterium Candidatus Chloracidobacterium thermophilum is the first aerobic chlorophototroph that has a type-I, homodimeric reaction center (RC). This organism and its type-I RCs were initially detected by the occurrence of pscA gene sequences, which encode the core subunit of the RC complex, in metagenomic sequence data derived from hot spring microbial mats. Here, we report the isolation and initial biochemical characterization of the type-I RC from Ca. C. thermophilum. After removal of chlorosomes, crude membranes were solubilized with 0.1% (w/v) n-dodecyl β-D-maltoside, and the RC complex was purified by ion-exchange chromatography. The RC complex comprised only two polypeptides: the reaction center core protein PscA and a 22-kDa carotenoid-binding protein denoted CbpC. The absorption spectrum showed a large, broad absorbance band centered at ∼483 nm from carotenoids as well as smaller Q(y) absorption bands at 672 and 812 nm from chlorophyll a and bacteriochlorophyll a, respectively. The light-induced difference spectra of whole cells, membranes, and the isolated RC showed maximal bleaching at 840 nm, which is attributed to the special pair and which we denote as P840. Making it unique among homodimeric type-I RCs, the isolated RC was photoactive in the presence of oxygen. Analyses by optical spectroscopy, chromatography, and mass spectrometry revealed that the RC complex contained 10.3 bacteriochlorophyll a(P), 6.4 chlorophyll a(PD), and 1.6 Zn-bacteriochlorophyll a(P)' molecules per P840 (12.8:8.0:2.0). The possible functions of the Zn-bacteriochlorophyll a(P)' molecules and the carotenoid-binding protein are discussed.

X-ray crystal structure of the light-independent protochlorophyllide reductase

Photosynthetic organisms adopt two different strategies for the reduction of the C17 = C18 double bond of protochlorophyllide (Pchlide) to form chlorophyllide a, the direct precursor of chlorophyll a (refs 1-4). The first involves the activity of the light-dependent Pchlide oxidoreductase, and the second involves the light-independent (dark-operative) Pchlide oxidoreductase (DPOR). DPOR is a nitrogenase-like enzyme consisting of two components, L-protein (a BchL dimer) and NB-protein (a BchN-BchB heterotetramer), which are structurally related to nitrogenase Fe protein and MoFe protein, respectively. Here we report the crystal structure of the NB-protein of DPOR from Rhodobacter capsulatus at a resolution of 2.3A. As expected, the overall structure is similar to that of nitrogenase MoFe protein: each catalytic BchN-BchB unit contains one Pchlide and one iron-sulphur cluster (NB-cluster) coordinated uniquely by one aspartate and three cysteines. Unique aspartate ligation is not necessarily needed for the cluster assembly but is essential for the catalytic activity. Specific Pchlide-binding accompanies the partial unwinding of an alpha-helix that belongs to the next catalytic BchN-BchB unit. We propose a unique trans-specific reduction mechanism in which the distorted C17-propionate of Pchlide and an aspartate from BchB serve as proton donors for C18 and C17 of Pchlide, respectively. Intriguingly, the spatial arrangement of the NB-cluster and Pchlide is almost identical to that of the P-cluster and FeMo-cofactor in nitrogenase MoFe-protein, illustrating that a common architecture exists to reduce chemically stable multibonds of porphyrin and dinitrogen.

Femtosecond Spectroscopy of the Primary Charge Separation in Reaction Centers ofChloroflexusaurantiacuswith Selective Excitation in the QYand Soret Bands

The primary charge separation and electron-transfer processes of photosynthesis occur in the reaction center (RC). Isolated RCs of the green filamentous anoxygenic phototrophic bacterium Chloroflexus aurantiacus were studied at room temperature by using femtosecond transient absorption spectroscopy with selective excitation. Upon excitation in the Q(Y) absorbance band of the bacteriochlorophyll (BChl) dimer (P) at 865 nm, a 7.0 +/- 0.5 ps kinetic component was observed in the 538 nm region (Q(X) band of the bacteriopheophytin (BPheo)), 750 nm region (Q(Y) band of the BPheo), and 920 nm region (stimulated emission of the excited-state of P), indicating that this lifetime represents electron transfer from P to BPheo. The same time constant was also observed upon 740 nm or 800 nm excitation. A longer lifetime (300 +/- 30 ps), which was assigned to the time of reduction of the primary quinone, Q(A), was also observed. The transient absorption spectra and kinetics all indicate that only one electron-transfer branch is involved in primary charge separation under these excitation conditions. However, the transient absorption changes upon excitation in the Soret band at 390 nm reveal a more complex set of energy and electron-transfer processes. By comparison to studies on the RCs of the purple bacterium Rhodobacter sphaeroides, we discuss the possible mechanism of electron-transfer pathway dependence on excitation energy and propose a model of the Cf. aurantiacus RC that better explains the observed results.

Analysis of absorption spectra of purple bacterial reaction centers in the near infrared region by higher order derivative spectroscopy

Reaction centers (RCs) of purple bacteria are uniquely suited objects to study the mechanisms of the photosynthetic conversion of light energy into chemical energy. A recently introduced method of higher order derivative spectroscopy [I.K. Mikhailyuk, H. Lokstein, A.P. Razjivin, A method of spectral subband decomposition by simultaneous fitting the initial spectrum and a set of its derivatives, J. Biochem. Biophys. Methods 63 (2005) 10–23] was used to analyze the NIR absorption spectra of RC preparations from Rhodobacter ( R.) sphaeroides strain 2R and Blastochloris ( B.) viridis strain KH, containing bacteriochlorophyll (BChl) a and b, respectively. Q y bands of individual RC porphyrin components (BChls and bacteriopheophytins, BPheo) were identified. The results indicate that the upper exciton level P y+ of the photo-active BChl dimer in RCs of R. sphaeroides has an absorption maximum of 810 nm. The blue shift of a complex integral band at approximately 800 nm upon oxidation of the RC is caused primarily by bleaching of P y+, rather than by an electrochromic shift of the absorption band(s) of the monomeric BChls. Likewise, the disappearance of a band peaking at 842 nm upon oxidation of RCs from B. viridis indicates that this band has to be assigned to P y+. A blue shift of an absorption band at approximately 830 nm upon oxidation of RCs of B. viridis is also essentially caused by the disappearance of P y+, rather than by an electrochromic shift of the absorption bands of monomeric BChls. Absorption maxima of the monomeric BChls, B B and B A are at 802 and 797 nm, respectively, in RCs of R. sphaeroides at room temperature. BPheo co-factors H B and H A peak at 748 and 758 nm, respectively, at room temperature. For B. viridis RCs the spectral positions of H B and H A were found to be 796 and 816 nm, respectively, at room temperature.

Lamellar Organization of Pigments in Chlorosomes, the Light Harvesting Complexes of Green Photosynthetic Bacteria

Chlorosomes of green photosynthetic bacteria constitute the most efficient light harvesting complexes found in nature. In addition, the chlorosome is the only known photosynthetic system where the majority of pigments (BChl) is not organized in pigment-protein complexes but instead is assembled into aggregates. Because of the unusual organization, the chlorosome structure has not been resolved and only models, in which BChl pigments were organized into large rods, were proposed on the basis of freeze-fracture electron microscopy and spectroscopic constraints. We have obtained the first high-resolution images of chlorosomes from the green sulfur bacterium Chlorobium tepidum by cryoelectron microscopy. Cryoelectron microscopy images revealed dense striations ∼20 Å apart. X-ray scattering from chlorosomes exhibited a feature with the same ∼20 Å spacing. No evidence for the rod models was obtained. The observed spacing and tilt-series cryoelectron microscopy projections are compatible with a lamellar model, in which BChl molecules aggregate into semicrystalline lateral arrays. The diffraction data further indicate that arrays are built from BChl dimers. The arrays form undulating lamellae, which, in turn, are held together by interdigitated esterifying alcohol tails, carotenoids, and lipids. The lamellar model is consistent with earlier spectroscopic data and provides insight into chlorosome self-assembly.

Light- and redox-dependent thermal stability of the reaction center of the photosynthetic Bacterium rhodobacter sphaeroides.

Irreversible loss of the photochemical activity and damage of the pigments (bacteriochlorophyll [Bchl] monomer, Bchl dimer [P] and bacteriopheophytin) by combined treatment with intense and continuous visible light and elevated temperature have been studied in a deoxygenated solution of reaction center (RC) protein from the nonsulfur purple photosynthetic bacterium Rhodobacter sphaeroides. Both the fraction of RC in the charge-separated redox state (P+Q-, where Q is a quinone electron acceptor) and the degradation of the pigments showed saturation as a function of increasing light intensity up to 400 mW cm(-2) (488/515 nm) or 1100 microE m(-2) s(-1) (white light). The thermal denaturation curves of the RC in the P+Q- redox state demonstrated broadening and 10-20 degrees C shift to lower temperature (after 30-90 min heat treatment) compared with those in the PQ redox state. Similar but less striking behavior was seen for RC of other redox states (P+Q and PQ-) generated either by light or by electrochemical treatment in the dark. These experiments suggest that it is not the intense light per se but the changes in the redox state of the protein that are responsible for the increased sensitivity to photo- and heat damage. The RC with a charge pair (P+Q-) is more vulnerable to elevated temperature than the RC with (P+Q or PQ-) or without (PQ) a single charge. To reveal both the thermodynamic and kinetic aspects of the denaturation, a simple three-state model of coupled reversible thermal and irreversible kinetic transitions is presented. These effects may have relevance to the heat stability of other redox proteins in bioenergetics.

Ab Initio Molecular Orbital Calculations of Electronic Couplings in the LH2 Bacterial Light-Harvesting Complex of Rps. Acidophila

The results of ab initio molecular orbital calculations of excited states and electronic couplings (for energy transfer) between the B800 and B850 bacteriochlorophyll a (Bchl) chromophores in the peripheral light-harvesting complex (LH2) of the purple photosynthetic bacterium Rhodopseudomonas acidophila are reported. Electronic couplings are estimated from “supermolecule” calculations of Bchl dimers using the CI-singles methodology and 3-21G* or 6-31G* basis sets. A scheme for dissecting the coupling into contributions from the Coulombic coupling and the short-range coupling (i.e., dependent on interchromophore orbital overlap) is reported. B850 couplings are calculated to be [total (Coulombic + short)]: intrapolypeptide dimer 320 (265 + 55) cm-1 and interpolypeptide dimer 255 (195 + 60) cm-1 at the CIS/6-31G* level. These results differ significantly from those estimated using the point dipole approximation. The effect of including Mg ligands (His residues) and H-bonding residues (Trp and Tyr) is also investigated. The consequences for superradiance and energy transfer dynamics and mechanism are discussed.

Electronic Excitations in Aggregates of Bacteriochlorophylls

Photosynthetic organisms have developed antenna systems to enlarge their cross section for capturing sunlight. These systems involve in some cases aggregates of bacteriochlorophylls (BChls). The structure of one such aggregate, a tightly coupled circular hexadecamer of BChls and a loosely coupled octamer of BChls, has recently been solved. In this paper we investigate the electronic excitations in these two aggregates as well as in monomeric BChl and BChl dimers by means of INDO/S-CI calculations. The results provide a detailed description of the properties of electronic states in the BChl aggregates (energies, dipole and transition dipole moments) that are relevant for the biological function of BChls, to absorb light and transfer energy on the subpicosecond time scale. The lower-energy excitations of the BChl hexadecamer are of exciton type, i.e., experiencing a strong coupling that leads to excitation delocalization over the entire aggregate. An effective Hamiltonian is provided which reproduces these ...

A comparative study of conserved protein interactions of the primary electron donor in photosynthetic purple bacterial reaction centers

The pigment–protein interactions within the binding site of the bacteriochlorophyll (BChl) dimer constituting the primary electron donor (P) in several, native, photosynthetic bacterial reaction centers have been determined using Fourier transform Raman spectroscopy. For reaction centers whose primary sequence data are available, and assuming a structural analogy with the Rb. sphaeroides RC whose high-resolution three-dimensional structure is known, amino acid residues donating hydrogen bonds to P are proposed. Consequently, one may propose the microenvironment structure of the primary donors studied and correlate this deduced structure with the known absorption and redox properties of the primary donors. In this ‘mini-review’ of our past work, we group and classify the primary donors with respect to their specific H-bonding interactions with the protein. This classification reveals trends in the H-bonding and certain physicochemical properties such as the P°/P⋅+ redox midpoint potential, the positive charge distribution over the dimeric primary donors in their oxidized radical cation state P⋅+, and the absorption maxima of the lower exciton Qy absorption band of P.