Type I photosynthetic reaction centres: structure and function

We review recent advances in the study of the photosystem I reaction centre, following the determination of a spectacular 2.5 A resolution crystal structure for this complex of Synechococcus elongatus. Photosystem I is proving different to type II reaction centres in structure and organization, and the mechanism of transmembrane electron transfer, and is providing insights into the control of function in reaction centres that operate at very low redox potentials. The photosystem I complex of oxygenic organisms has a counterpart in non-oxygenic bacteria, the strictly anaerobic phototrophic green sulphur bacteria and heliobacteria. The most distinctive feature of these type I reaction centres is that they contain two copies of a large core polypeptide (i.e. a homodimer), rather than a heterodimeric arrangement of two related, but different, polypeptides as in the photosystem I complex. To compare the structural organization of the two forms of type I reaction centre, we have modelled the structure of the central region of the reaction centre from green sulphur bacteria, using sequence alignments and the structural coordinates of the S. elongatus Photosystem I complex. The outcome of these modelling studies is described, concentrating on regions of the type I reaction centre where important structure-function relationships have been demonstrated or inferred.     

Photosynthetic reaction centres: variations on a common structural theme?

From their hybrid properties, the reaction centres of green sulphur bacteria and heliobacteria seem to be the missing links between the two branches of the reaction centre family, typified by higher plant photosystem I and the purple bacterial reaction centre. This suggests that all of the diverse types of photosynthetic reaction centres have closer structural resemblances than was previously thought.     

Insights into the evolution of the antenna domains of Type-I and Type-II photosynthetic reaction centres through homology modelling

The (bacterio)chlorophylls of photosynthetic antenna and reaction centre complexes are bound to the protein via a fifth, axial ligand to the central magnesium atom. A number of the amino acids identified as providing such ligands are conserved between the large antenna of the cyanobacterial Type-I reaction centre and smaller antennas of the Type-I reaction centres of green sulphur bacteria and heliobacteria, and these numbers match closely the estimated number of antenna bacteriochlorophylls in the latter. The possible organisation of the antenna in the latter reaction centres is discussed, as is the mechanism by which the more pigment-rich antenna of the cyanobacterial reaction centre evolved. The homology modelling approach is also extended to the six-helix antenna proteins CP47 and CP43 associated with the Photosystem II reaction centre.     

Photochemically induced dynamic nuclear polarization in the reaction center of the green sulphur bacterium Chlorobium tepidum observed by 13C MAS NMR

Photochemically induced dynamic nuclear polarization has been observed in reaction centres of the green sulphur bacterium Chlorobium tepidum by (13)C magic-angle spinning solid-state NMR under continuous illumination with white light. An almost complete set of chemical shifts of the aromatic ring carbons of a BChl a molecule has been obtained. All light-induced (13)C NMR signals appear to be emissive, which is similar to the pattern observed in the reaction centers of plant photosystem I and purple bacterial reaction centres of Rhodobacter sphaeroides wild type. The donor in RCs of green sulfur bacteria clearly differs from the substantially asymmetric special pair of purple bacteria and appears to be similar to the more symmetric donor of photosystem I.     

Phylogenetic Analyses of the Core Antenna Domain: Investigatingthe Origin of Photosystem I

Phototrophy, the conversion of light to biochemical energy, occurs throughout the Bacteria and plants, however, debate continues over how different phototrophic mechanisms and the bacteria that contain them are related. There are two types of phototrophic mechanisms in the Bacteria: reaction center type 1 (RC1) has core and core antenna domains that are parts of a single polypeptide, whereas reaction center type 2 (RC2) is composed of short core proteins without antenna domains. In cyanobacteria, RC2 is associated with separate core antenna proteins that are homologous to the core antenna domains of RC1. We reconstructed evolutionary relationships among phototrophic mechanisms based on a phylogeny of core antenna domains/proteins. Core antenna domains of 46 polypeptides were aligned, including the RC1 core proteins of heliobacteria, green sulfur bacteria, and photosystem I (PSI) of cyanobacteria and plastids, plus core antenna proteins of photosystem II (PSII) from cyanobacteria and plastids. Maximum likelihood, parsimony, and neighbor joining methods all supported a single phylogeny in which PSII core antenna proteins (PsbC, PsbB) arose within the cyanobacteria from duplications of the RC1-associated core antenna domains and accessory antenna proteins (IsiA, PcbA, PcbC) arose from duplications of PsbB. The data indicate an evolutionary history of RC1 in which an initially homodimeric reaction center was vertically transmitted to green sulfur bacteria, heliobacteria, and an ancestor of cyanobacteria. A heterodimeric RC1 (=PSI) then arose within the cyanobacterial lineage. In this scenario, the current diversity of core antenna domains/proteins is explained without a need to invoke horizontal transfer.This article contains online-only supplementary material.Reviewing Editor: Dr. W. Ford Doolittle     

The reaction centre from green sulphur bacteria: progress towards structural elucidation

The reaction centre (RC) of green sulphur bacteria is a FeS-type RC, as are the RCs of Photosystems I (PS I) of oxygenic photosynthetic organisms and of heliobacteria. The core domains of both green sulphur bacterial and heliobacterial RCs are considered to be homodimeric, in contrast to those of purple bacteria, PS I and Photosystem II (PS II). This paper briefly describes the techniques of electron microscopy and image processing suited to investigate the structure of these proteins. Recent advances in the study of the structure of the green sulphur bacterial RC, primarily achieved by the application of scanning transmission electron microscopy, are reviewed.This revised version was published online in October 2005 with corrections to the Cover Date.     

Photochemically induced dynamic nuclear polarization in the reaction center of the green sulphur bacterium Chlorobium tepidum observed by C MAS NMR

Photochemically induced dynamic nuclear polarization has been observed in reaction centres of the green sulphur bacterium Chlorobium tepidum by ¹³C magic-angle spinning solid-state NMR under continuous illumination with white light. An almost complete set of chemical shifts of the aromatic ring carbons of a BChl a molecule has been obtained. All light-induced ¹³C NMR signals appear to be emissive, which is similar to the pattern observed in the reaction centers of plant photosystem I and purple bacterial reaction centres of Rhodobacter sphaeroides wild type. The donor in RCs of green sulfur bacteria clearly differs from the substantially asymmetric special pair of purple bacteria and appears to be similar to the more symmetric donor of photosystem I.     

The crystal structure of an asymmetric complex of the two nucleotide binding components of proton-translocating transhydrogenase

BACKGROUND: Membrane-bound ion translocators have important functions in biology, but their mechanisms of action are often poorly understood. Transhydrogenase, found in animal mitochondria and bacteria, links the redox reaction between NAD(H) and NADP(H) to proton translocation across a membrane. Linkage is achieved through changes in protein conformation at the nucleotide binding sites. The redox reaction takes place between two protein components located on the membrane surface: dI, which binds NAD(H), and dIII, which binds NADP(H). A third component, dII, provides a proton channel through the membrane. Intact membrane-located transhydrogenase is probably a dimer (two copies each of dI, dII, and dIII). RESULTS: We have solved the high-resolution crystal structure of a dI:dIII complex of transhydrogenase from Rhodospirillum rubrum-the first from a transhydrogenase of any species. It is a heterotrimer, having two polypeptides of dI and one of dIII. The dI polypeptides fold into a dimer. The loop on dIII, which binds the nicotinamide ring of NADP(H), is inserted into the NAD(H) binding cleft of one of the dI polypeptides. The cleft of the other dI is not occupied by a corresponding dIII component. CONCLUSIONS: The redox step in the transhydrogenase reaction is readily visualized; the NC4 atoms of the nicotinamide rings of the bound nucleotides are brought together to facilitate direct hydride transfer with A-B stereochemistry. The asymmetry of the dI:dIII complex suggests that in the intact enzyme there is an alternation of conformation at the catalytic sites associated with changes in nucleotide binding during proton translocation.     

Characterisation of the PsbX protein from Photosystem II and light regulation of its gene expression in higher plants

The location and expression of the previously uncharacterised photosystem II subunit PsbX have been analysed in higher plants. We show that this protein is a component of photosystem II (PSII) core particles but absent from light-harvesting complexes or PSII reaction centres. PsbX is, however, localised to the near vicinity of the reaction centre because it can be cross-linked to cytochrome b559, which is known to be associated with the D1/D2 dimer. We also show that the expression of this protein is tightly regulated by light, since neither protein nor mRNA is found in dark-grown plants.     

Evolution of heliobacteria: Implications for photosynthetic reaction center complexes

The evolutionary position of the heliobacteria, a group of green photosynthetic bacteria with a photosynthetic apparatus functionally resembling Photosystem I of plants and cyanobacteria, has been investigated with respect to the evolutionary relationship to Gram-positive bacteria and cyanobacteria. On the basis of 16S rRNA sequence analysis, the heliobacteria appear to be most closely related to Gram-positive bacteria, but also an evolutionary link to cyanobacteria is evident. Interestingly, a 46-residue domain including the putative sixth membrane-spanning region of the heliobacterial reaction center protein shows rather strong similarity (33% identity and 72% similarity) to a region including the sixth membrane-spanning region of the CP47 protein, a chlorophyll-binding core antenna polypeptide of Photosystem II. The N-terminal half of the heliobacterial reaction center polypeptide shows a moderate sequence similarity (22% identity over 232 residues) with the CP47 protein, which is significantly more than the similarity with the Photosystem I core polypeptides in this region. An evolutionary model for photosynthetic reaction center complexes is discussed, in which an ancestral homodimeric reaction center protein (possibly resembling the heliobacterial reaction center protein) with 11 membrane-spanning regions per polypeptide has diverged to give rise to core of Photosystem I, Photosystem II, and of the photosynthetic apparatus in green, purple, and heliobacteria.     

Fourier transform infrared spectroscopy of primary electron donors in type I photosynthetic reaction centers.

The vibrational properties of the primary electron donors (P) of type I photosynthetic reaction centers, as investigated by Fourier transform infrared (FTIR) difference spectroscopy in the last 15 years, are briefly reviewed. The results obtained on the microenvironment of the chlorophyll molecules in P700 of photosystem I and of the bacteriochlorophyll molecules in P840 of the green bacteria (Chlorobium) and in P798 of heliobacteria are presented and discussed with special attention to the bonding interactions with the protein of the carbonyl groups and of the central Mg atom of the pigments. The observation of broad electronic transitions in the mid-IR for the cationic state of all the primary donors investigated provides evidence for charge repartition over two (B)Chl molecules. In the green sulfur bacteria and the heliobacteria, the assignments proposed for the carbonyl groups of P and P(+) are still very tentative. In contrast, the axial ligands of P700 in photosystem I have been identified and the vibrational properties of the chlorophyll (Chl) molecules involved in P700, P700(+), and (3)P700 are well described in terms of two molecules, denoted P(1) and P(2), with very different hydrogen bonding patterns. While P(1) has hydrogen bonds to both the 9-keto and the 10a-ester C=O groups and bears all the triplet character in (3)P700, the carbonyl groups of P(2) are free from hydrogen bonding. The positive charge in P700(+) is shared between the two Chl molecules with a ratio ranging from 1:1 to 2:1, in favor of P(2), depending on the temperature and the species. The localization of the triplet in (3)P700 and of the unpaired electron in P700(+) deduced from FTIR spectroscopy is in sharp contrast with that resulting from the analysis of the magnetic resonance experiments. However, the FTIR results are in excellent agreement with the most recent structural model derived from X-ray crystallography of photosystem I at 2.5 A resolution that reveals the hydrogen bonds to the carbonyl groups of the Chl in P700 as well as the histidine ligands of the central Mg atoms predicted from the FTIR data.     

Fourier transform infrared spectroscopy of special pair bacteriochlorophylls in homodimeric reaction centers of heliobacteria and green sulfur bacteria

Heliobacteria and green sulfur bacteria have type I homodimeric reaction centers analogous to photosystem I. One remaining question regarding these homodimeric reaction centers is whether the structures and electron transfer reactions are truly symmetric or not. This question is relevant to the origin of the heterodimeric reaction centers, such as photosystem I and type II reaction centers. In this mini-review, Fourier transform infrared studies on the special pair bacteriochlorophylls, P798 in heliobacteria and P840 in green sulfur bacteria, are summarized. The data are reinterpreted in the light of the X-ray crystallographic structure of photosystem I and the sequence alignments of type I reaction center proteins, and discussed in terms of hydrogen bonding interactions and the symmetry of charge distribution over the dimer.     

Thinking About the Evolution of Photosynthesis

Photosynthesis is an ancient process on Earth. Chemical evidence and recent fossil finds indicate that cyanobacteria existed 2.5-2.6 billion years (Ga) ago, and these were certainly preceded by a variety of forms of anoxygenic photosynthetic bacteria. Carbon isotope data suggest autotrophic carbon fixation was taking place at least a billion years earlier. However, the nature of the earliest photosynthetic organisms is not well understood. The major elements of the photosynthetic apparatus are the reaction centers, antenna complexes, electron transfer complexes and carbon fixation machinery. These parts almost certainly have not had the same evolutionary history in all organisms, so that the photosynthetic apparatus is best viewed as a mosaic made up of a number of substructures each with its own unique evolutionary history. There are two schools of thought concerning the origin of reaction centers and photosynthesis. One school pictures the evolution of reaction centers beginning in the prebiotic phase while the other school sees reaction centers evolving later from cytochrome b in bacteria. Two models have been put forth for the subsequent evolution of reaction centers in proteobacteria, green filamentous (non-sulfur) bacteria, cyanobacteria, heliobacteria and green sulfur bacteria. In the selective loss model the most recent common ancestor of all subsequent photosynthetic systems is postulated to have contained both RC1 and RC2. The evolution of reaction centers in proteobacteria and green filamentous bacteria resulted from the loss of RC1, while the evolution of reaction centers in heliobacteria and green sulfur bacteria resulted from the loss of RC2. Both RC1 and RC2 were retained in the cyanobacteria. In the fusion model the most recent common ancestor is postulated to have given rise to two lines, one containing RC1 and the other containing RC2. The RC1 line gave rise to the reaction centers of heliobacteria and green sulfur bacteria, and the RC2 line led to the reaction centers of proteobacteria and green filamentous bacteria. The two reaction centers of cyanobacteria were the result of a genetic fusion of an organism containing RC1 and an organism containing RC2. The evolutionary histories of the various classes of antenna/light-harvesting complexes appear to be completely independent. The transition from anoxygenic to oxygenic photosynthesis took place when the cyanobacteria learned how to use water as an electron donor for carbon dioxide reduction. Before that time hydrogen peroxide may have served as a transitional donor, and before that, ferrous iron may have been the original source of reducing power.     

Characterization of new phototrophic heliobacteria and their habitats

Specific enrichment culture methods were used to isolate new phototrophic heliobacteria (anoxygenic phototrophic bacteria containing bacteriochlorophyll g) from various natural samples. A survey of terrestrial and aquatic habitats yielded heliobacteria only from soils, in particular rice soils, and from certain hot springs. Thirteen nonthermophilic and 7 thermophilic (capable of growth above 50) strains of heliobacteria were isolated from such habitats and characterized as to their basic cellular and nutritional properties. Phylogenetic studies of four strains showed them to be related to known species of heliobacteria. It is concluded that, unlike phototrophic purple and green bacteria, heliobacteria are primarily (if not exclusively) terrestrial, except for hot spring species. This suggests that the ecology of heliobacteria is fundamentally different from that of other anoxyphototrophs.     

Design of a redox-linked active metal site: manganese bound to bacterial reaction centers at a site resembling that of photosystem II

Metals bound to proteins perform a number of crucial biological reactions, including the oxidation of water by a manganese cluster in photosystem II. Although evolutionarily related to photosystem II, bacterial reaction centers lack both a strong oxidant and a manganese cluster for mediating the multielectron and proton transfer needed for water oxidation. In this study, carboxylate residues were introduced by mutagenesis into highly oxidizing reaction centers at a site homologous to the manganese-binding site of photosystem II. In the presence of manganese, light-minus-dark difference optical spectra of reaction centers from the mutants showed a lack of the oxidized bacteriochlorophyll dimer, while the reduced primary quinone was still present, demonstrating that manganese was serving as a secondary electron donor. On the basis of these steady-state optical measurements, the mutant with the highest-affinity site had a dissociation constant of approximately 1 microM. For the highest-affinity mutant, a first-order rate with a lifetime of 12 ms was observed for the reduction of the oxidized bacteriochlorophyll dimer by the bound manganese upon exposure to light. The dependence of the amplitude of this component on manganese concentration yielded a dissociation constant of approximately 1 muM, similar to that observed in the steady-state measurements. The three-dimensional structure determined by X-ray diffraction of the mutant with the high-affinity site showed that the binding site contains a single bound manganese ion, three carboxylate groups (including two groups introduced by mutagenesis), a histidine residue, and a bound water molecule. These reaction centers illustrate the successful design of a redox active metal center in a protein complex.     

Excitation pressure regulates the activation energy for recombination events in the photosystem II reaction centres of Chlamydomonas reinhardtii.

Using in vivo thermoluminescence, we examined the effects of growth irradiance and growth temperature on charge recombination events in photosystem II reaction centres of the model green alga Chlamydomonas reinhardtii. We report that growth at increasing irradiance at either 29 or 15 degrees C resulted in comparable downward shifts in the temperature peak maxima (T(M)) for S2QB- charge pair recombination events, with minimal changes in S2QA- recombination events. This indicates that such growth conditions decrease the activation energy required for S2QB- charge pair recombination events with no concomitant change in the activation energy for S2QA- recombination events. This resulted in a decrease in the DeltaT(M) between S2QA- and S2QB- recombination events, which was reversible when shifting cells from low to high irradiance and back to low irradiance at 29 degrees C. We interpret these results to indicate that the redox potential of QB was modulated independently of QA, which consequently narrowed the redox potential gap between QA and QB in photosystem II reaction centres. Since a decrease in the DeltaT(M) between S2QA- and S2QB- recombination events correlated with growth at increasing excitation pressure, we conclude that acclimation to growth under high excitation pressure narrows the redox potential gap between QA and QB in photosystem II reaction centres, enhancing the probability for reaction center quenching in C. reinhardtii. We discuss the molecular basis for the modulation of the redox state of QB, and suggest that the potential for reaction center quenching complements antenna quenching via the xanthophyll cycle in the photoprotection of C. reinhardtii from excess light.     

Distribution of chloropigments in suspended particulate matter and benthic microbial mat of a meromictic lake, Lake Kaiike, Japan

We investigated the distribution of chloropigments in a small meromictic lake, Lake Kaiike, south-west Japan. In the water-column, concentrations of Chl a related to cyanobacteria, BChl a related to purple sulphur bacteria, and three types of BChl e homologues (BChls e1, e2 and e3) related to brown-coloured green sulphur bacteria, were maximal at the redox boundary. Below the redox boundary, absolute concentrations of Chl a and BChl a gradually decreased with depth, whereas BChls e remained rather constant. Suspended particulate matter (SPM) at the deeper region of the anoxic water-column was enriched in highly alkylated BChl e homologues compared with SPM at the redox boundary. The shift in the relative content of highly alkylated BChl e homologues beneath the boundary was associated with community related adaptation of brown-coloured green sulphur bacteria to changes in light quality/quantity, resulting from the optical absorption and reflectance of SPMs in the overlying water-column. Benthic microbial mats were characterized by high abundances of BChls e, in which highly alkylated homologues were substantially abundant. This suggests that the BChls e in the microbial mat may be derived from the low-light adapted brown-coloured green sulphur bacteria forming the bacterial mat.     

The oxidation state of the photosystem II manganese cluster influences the structure of manganese stabilizing protein

Exposure of photosystem II membranes to trypsin that has been treated to inhibit chymotrypsin activity produces limited hydrolysis of manganese stabilizing protein. Exposure to chymotrypsin under the same conditions yields substantial digestion of the protein. Further probing of the unusual insensitivity of manganese stabilizing protein to trypsin hydrolysis reveals that increasing the temperature from 4 to 25 degrees C will cause some acceleration in the rate of proteolysis. However, addition of low (100 microM) concentrations of NH2OH, that are sufficient to reduce, but not destroy, the photosystem II Mn cluster, causes a change in PS II-bound manganese stabilizing protein that causes it to be rapidly digested by trypsin. Immunoblot analyses with polyclonal antibodies directed against the N-terminus of the protein, or against the entire sequence show that trypsin cleavage produces two distinct peptide fragments estimated to be in the 17-20 kDa range, consistent with proposals that there are 2 mol of the protein/mol photosystem II. The correlation of trypsin sensitivity with Mn redox state(s) in photosystem II suggest that manganese stabilizing protein may interact either directly with Mn, or alternatively, that the polypeptide is bound to another protein of the photosystem II reaction center that is intimately involved in binding and redox activity of Mn.     

Daddy, where did (PS)I come from?

The reacton centre I (RCI)-type photosystems from plants, cyano-, helio- and green sulphur bacteria are compared and the essential properties of an archetypal RCI are deduced. Species containing RCI-type photosystems most probably cluster together on a common branch of the phylogenetic tree. The predicted branching order is green sulphur, helio- and cyanobacteria. Striking similarities between RCI- and RCII-type photosystems recently became apparent in the three-dimensional structures of photosystem I (PSI), PSII and RCII. The phylogenetic relationship between all presently known photosystems is analysed suggesting (a) RCI as the ancestral photosystem and (b) the descendence of PSII from RCI via gene duplication and gene splitting. An evolutionary model trying to rationalise available data is presented.