Bacterial Photosynthesis Research Articles

Photobiological transformation of hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) using Rhodobacter sphaeroides

Pump-and-treat strategies for groundwater containing explosives may be necessary when the contaminated water approaches sensitive receptors. This project investigated bacterial photosynthesis as a strategy for ex situ treatment, using light as the primary energy source to facilitate RDX transformation. The objective was to characterize the ability of photosynthetic Rhodobacter sphaeroides (strain ATCC® 17023 ™) to transform the high-energy explosive RDX. R. sphaeroides transformed 30 μM RDX within 40 h under light conditions; RDX was not fully transformed in the dark (non-photosynthetic conditions), suggesting that photosynthetic electron transfer was the primary mechanism. Experiments with RDX demonstrated that succinate and malate were the most effective electron donors for photosynthesis, but glycerol was also utilized as a photosynthetic electron donor. RDX was transformed irrespective of the presence of carbon dioxide. The electron shuttling compound anthraquinone-2,6-disulfonate (AQDS) increased transformation kinetics in the absence of CO2, when the cells had excess NADPH that needed to be re-oxidized because there was limited CO2 for carbon fixation. When CO2 was added, the cells generated more biomass, and AQDS had no stimulatory effect. End products indicated that RDX carbon became CO2, biomass, and a soluble, uncharacterized aqueous metabolite, determined using 14C-labeled RDX. These data are the first to suggest that photobiological explosives transformation is possible and will provide a framework for which phototrophy can be used in environmental restoration of explosives contaminated water.

New Dimensions in Microbial Ecology-Functional Genes in Studies to Unravel the Biodiversity and Role of Functional Microbial Groups in the Environment.

During the past decades, tremendous advances have been made in the possibilities to study the diversity of microbial communities in the environment. The development of methods to study these communities on the basis of 16S rRNA gene sequences analysis was a first step into the molecular analysis of environmental communities and the study of biodiversity in natural habitats. A new dimension in this field was reached with the introduction of functional genes of ecological importance and the establishment of genetic tools to study the diversity of functional microbial groups and their responses to environmental factors. Functional gene approaches are excellent tools to study the diversity of a particular function and to demonstrate changes in the composition of prokaryote communities contributing to this function. The phylogeny of many functional genes largely correlates with that of the 16S rRNA gene, and microbial species may be identified on the basis of functional gene sequences. Functional genes are perfectly suited to link culture-based microbiological work with environmental molecular genetic studies. In this review, the development of functional gene studies in environmental microbiology is highlighted with examples of genes relevant for important ecophysiological functions. Examples are presented for bacterial photosynthesis and two types of anoxygenic phototrophic bacteria, with genes of the Fenna-Matthews-Olson-protein (fmoA) as target for the green sulfur bacteria and of two reaction center proteins (pufLM) for the phototrophic purple bacteria, with genes of adenosine-5′phosphosulfate (APS) reductase (aprA), sulfate thioesterase (soxB) and dissimilatory sulfite reductase (dsrAB) for sulfur oxidizing and sulfate reducing bacteria, with genes of ammonia monooxygenase (amoA) for nitrifying/ammonia-oxidizing bacteria, with genes of particulate nitrate reductase and nitrite reductases (narH/G, nirS, nirK) for denitrifying bacteria and with genes of methane monooxygenase (pmoA) for methane oxidizing bacteria.

The Path to Thioredoxin and Redox Regulation in Chloroplasts.

After a brief discussion of my graduate work at Duke University, I describe a series of investigations on redox proteins at the University of California, Berkeley. Starting with ferredoxin from fermentative bacteria, the Berkeley research fostered experiments that uncovered a pathway for fixing CO2 in bacterial photosynthesis. The carbon work, in turn, opened new vistas, including the discovery that thioredoxin functions universally in regulating the Calvin-Benson cycle in oxygenic photosynthesis. These experiments, which took place over a 50-year period, led to the formulation of a set of biological principles and set the stage for research demonstrating a role for redox in the regulation of previously unrecognized processes extending far beyond photosynthesis.

Exploring the effects of photon correlations from thermal sources on bacterial photosynthesis

Thermal light sources can produce photons with strong spatial correlations. We study the role that these correlations might potentially play in bacterial photosynthesis. Our findings show a relationship between the transversal distance between consecutive absorptions and the efficiency of the photosynthetic process. Furthermore, membranes where the clustering of core complexes (so-called RC-LH1) is high, display a range where the organism profits maximally from the spatial correlation of the incoming light. By contrast, no maximum is found for membranes with low core-core clustering. We employ a detailed membrane model with state-of-the-art empirical inputs. Our results suggest that the organization of the membrane’s antenna complexes may be well-suited to the spatial correlations present in an natural light source. Future experiments will be needed to test this prediction.

Unified analysis of ensemble and single-complex optical spectral data from light-harvesting complex-2 chromoproteins for gaining deeper insight into bacterial photosynthesis.

Bacterial light-harvesting pigment-protein complexes are very efficient at converting photons into excitons and transferring them to reaction centers, where the energy is stored in a chemical form. Optical properties of the complexes are known to change significantly in time and also vary from one complex to another; therefore, a detailed understanding of the variations on the level of single complexes and how they accumulate into effects that can be seen on the macroscopic scale is required. While experimental and theoretical methods exist to study the spectral properties of light-harvesting complexes on both individual complex and bulk ensemble levels, they have been developed largely independently of each other. To fill this gap, we simultaneously analyze experimental low-temperature single-complex and bulk ensemble optical spectra of the light-harvesting complex-2 (LH2) chromoproteins from the photosynthetic bacterium Rhodopseudomonas acidophila in order to find a unique theoretical model consistent with both experimental situations. The model, which satisfies most of the observations, combines strong exciton-phonon coupling with significant disorder, characteristic of the proteins. We establish a detailed disorder model that, in addition to containing a C_{2}-symmetrical modulation of the site energies, distinguishes between static intercomplex and slow conformational intracomplex disorders. The model evaluations also verify that, despite best efforts, the single-LH2-complex measurements performed so far may be biased toward complexes with higher Huang-Rhys factors.

Elaboration of an unexplored substitution site in synthetic bacteriochlorins

The ability to introduce substituents at designated sites about the perimeter of synthetic bacteriochlorins – analogs of bacteriochlorophylls of bacterial photosynthesis – remains a subject of ongoing study. Here, the self-condensation of a dihydrodipyrrin-dioxolane affords a 5-[2-(trimethylsiloxy)ethoxy]bacteriochlorin. Like a 5-methoxybacteriochlorin, the latter undergoes regioselective bromination at the 15-position, directed by the distal 5-alkoxy group. On the other hand, attempted bromination of a bacteriochlorin bearing a 5-(2-hydroxyethoxy) group resulted in intramolecular ether formation with the adjacent β-pyrroline position to give an annulated dioxepine ring (confirmed by single-crystal X-ray structural analysis). The hydroxyethoxy group at the 5-position can be derivatized by acylation. In addition, the installation of auxochromes (methoxycarbonyl, phenylethynyl) at the β-pyrrole rings causes a substantial bathochromic shift of the long-wavelength absorption band (812 nm) and companion fluorescence emission band (821 nm). Taken together, the modification of the 5-substituent complements existing methods for installing a single substituent on the bacteriochlorin macrocycle.

Communication: Microsecond dynamics of the protein and water affect electron transfer in a bacterial bc(1) complex.

Cross-membrane electron transport between cofactors localized in proteins of mitochondrial respiration and bacterial photosynthesis is the source of all biological energy. The statistics and dynamics of nuclear fluctuations in these protein/membrane/water heterogeneous systems are critical for their energetic efficiency. The results of 13 μs of atomistic molecular dynamics simulations of the membrane-bound bc1 bacterial complex are analyzed here. The reaction is affected by a broad spectrum of nuclear modes, with the slowest dynamics in the range of time-scales ∼0.1-1.6 μs contributing half of the reaction reorganization energy. Two reorganization energies are required to describe protein electron transfer due to dynamical arrest of protein conformations on the observation window. This mechanistic distinction allows significant lowering of activation barriers for reactions in proteins.

Energy efficiency of the sunlight harvesting and storing system in bacterial photosynthesis: comparison with semiconductor photovoltaic cells

The light phase of photosynthesis is considered a joint operation of 2 functional pigment-protein complexes: a light harvesting antenna, absorbing sunlight in a wide spectral range, and a reaction center, utilizing the energy of absorbed light quanta in photochemical charge separation reactions. These complexes allow converting solar energy to the energy of specific biomolecules with high quantum efficiency. However, before being transferred to reaction centers, the solar energy is stored in the lowest excited state of pigment molecules of the light harvesting antenna that partially convert light quantum energy into heat. These energy losses bring a significant reduction of energy efficiency of photosynthesis in view of a very wide spectral range of photosynthetically active sunlight. In the current study we analyzed the energy efficiency of sunlight harvesting and storing in different photosynthetic bacteria with different absorption bands. We showed that simultaneous exploitation of several such photosynthetic organisms leads to an increased total energy efficiency in terms of harvesting sunlight of the wider spectra. Maximal values of energy efficiencies of the sunlight harvesting and storing system in photosynthetic bacteria and semiconductor photovoltaic cells are compared, and perspectives on practical use of photosynthetic bacteria as solar energy converters are discussed.

Survivability of Photosynthetic Bacteria In Non-Terrestrial Light

We explore whether a basic process of Life on Earth. Bacterial photosynthesis can in principle survive under alien light conditions defined by abnormal temporal correlations in the incident photon absorption times. Though unlike radiation from our own Sun, such extreme photon statistics have already been demonstrated under laboratory conditions and hence are allowed by the laws of physics. Our analysis exploits a detailed membrane model of the bacterial photosynthetic system Rs. Photometricum using state-of-the-art empirical inputs. Our results show that a broad range of extreme light conditions including those far beyond terrestrial sunlight do indeed support the metabolic needs of the terrestrial bacteria.

Regulation of nitrite resistance of the cytochrome cbb3 oxidase by cytochrome c ScyA in Shewanella oneidensis.

Cytochrome c proteins, as enzymes to exchange electrons with substrates or as pure electron carriers to shuttle electrons, play vital roles in bacterial respiration and photosynthesis. In Shewanella oneidensis, a research model for the respiratory diversity, at least 42 c-type cytochromes are predicted to be encoded in the genome and are regarded to be the foundation of its highly branched electron transport pathways. However, only a small number of c-type cytochromes have been extensively studied. In this study, we identify soluble cytochrome c ScyA as an important factor influencing the nitrite resistance of a strain devoid of the bd oxidase by utilizing a newly developed transposon mutagenesis vector, which enables overexpression of the gene(s) downstream of the insertion site. We show that when in overabundance ScyA facilitates growth against nitrite inhibition by enhancing nitrite resistance of the cbb3 oxidase. Based on the data presented in this study, we suggest two possible mechanisms underlying the observed effect of ScyA: (1) ScyA increases electron flow to the cbb3 oxidase; (2) ScyA promotes nitrite resistance of the cbb3 oxidase, possibly by direct interaction.

Assembly of functional photosystem complexes in Rhodobacter sphaeroides incorporating carotenoids from the spirilloxanthin pathway

Carotenoids protect the photosynthetic apparatus against harmful radicals arising from the presence of both light and oxygen. They also act as accessory pigments for harvesting solar energy, and are required for stable assembly of many light-harvesting complexes. In the phototrophic bacterium Rhodobacter (Rba.) sphaeroides phytoene desaturase (CrtI) catalyses three sequential desaturations of the colourless carotenoid phytoene, extending the number of conjugated carbon–carbon double bonds, N, from three to nine and producing the yellow carotenoid neurosporene; subsequent modifications produce the yellow/red carotenoids spheroidene/spheroidenone (N=10/11). Genomic crtI replacements were used to swap the native three-step Rba. sphaeroides CrtI for the four-step Pantoea agglomerans enzyme, which re-routed carotenoid biosynthesis and culminated in the production of 2,2′-diketo-spirilloxanthin under semi-aerobic conditions. The new carotenoid pathway was elucidated using a combination of HPLC and mass spectrometry. Premature termination of this new pathway by inactivating crtC or crtD produced strains with lycopene or rhodopin as major carotenoids. All of the spirilloxanthin series carotenoids are accepted by the assembly pathways for LH2 and RC–LH1–PufX complexes. The efficiency of carotenoid-to-bacteriochlorophyll energy transfer for 2,2′-diketo-spirilloxanthin (15 conjugated CC bonds; N=15) in LH2 complexes is low, at 35%. High energy transfer efficiencies were obtained for neurosporene (N=9; 94%), spheroidene (N=10; 96%) and spheroidenone (N=11; 95%), whereas intermediate values were measured for lycopene (N=11; 64%), rhodopin (N=11; 62%) and spirilloxanthin (N=13; 39%). The variety and stability of these novel Rba. sphaeroides antenna complexes make them useful experimental models for investigating the energy transfer dynamics of carotenoids in bacterial photosynthesis.

Cyanobacteria as an Experimental Platform for Modifying Bacterial and Plant Photosynthesis.

OPINION article Front. Bioeng. Biotechnol., 21 April 2014Sec. Synthetic Biology https://doi.org/10.3389/fbioe.2014.00007

Apo-bacteriophytochromes modulate bacterial photosynthesis in response to low light

Bacteriophytochromes (BphPs) are light-sensing regulatory proteins encoded by photosynthetic and nonphotosynthetic bacteria. This protein class has been characterized structurally, but its biological activities remain relatively unexplored. Two BphPs in the anoxygenic photosynthetic bacterium Rhodopseudomonas palustris, designated regulatory proteins RpBphP2 and RpBphP3, are configured as light-regulated histidine kinases, which initiate a signal transduction system that controls expression of genes for the low light harvesting 4 (LH4) antenna complex. In vitro, RpBphP2 and RpBphP3 respond to light quality by reversible photoconversion, a property that requires the light-absorbing chromophore biliverdin. In vivo, RpBphP2 and RpBphP3 are both required for the expression of the LH4 antenna complex under anaerobic conditions, but biliverdin requires oxygen for its synthesis by heme oxygenase. On further investigation, we found that the apo-bacteriophytochrome forms of RpBphP2 and RpBphP3 are necessary and sufficient to control LH4 expression in response to light intensity in conjunction with other signal transduction proteins. One possibility is that the system senses a reduced quinone pool generated when light energy is absorbed by bacteriochlorophyll. The biliverdin-bound forms of the BphPs have the additional property of being able to fine-tune LH4 expression in response to light quality. These observations support the concept that some bacteriophytochromes can function with or without a chromophore and may be involved in regulating physiological processes not directly related to light sensing.

Special Issue of Photosynthetic Research

This Special Issue of Photosynthesis Research honors Louis M. N. Duysens, Roderick K. Clayton, and George Feher, three pioneering researchers whose work on bacterial photosynthesis laid much of the groundwork for our understanding of the role of the reaction center in photosynthetic light energy conversion. Their key discoveries are briefly summarized and an overview of the special issue is presented.

Modeling of reversible charge separation in reaction centers of photosynthesis: An incoherent approach

Primary charge separation in reaction centers (RCs) of bacterial photosynthesis is modeled in this work. An incoherent population dynamics of RCs states is formulated by kinetic equations. It is assumed that charge separation is accompanied by regular motion of the system along additional coordinates. This motion modulates an energetics of the reactions, and this modulation causes femtosecond oscillations in the population of the states. The best qualitative and quantitative accordance with experimental data on native, modified and mutant RCs of Rba. sphaeroides is achieved in the five states model that includes two excited states P⁎905BAHA and P⁎940BAHA and three charge separated states I, P+BA−HA and P+BAHA− (P is a primary electron donor, bacteriochlorophyll dimer, BA and HA are electron acceptors, monomeric bacteriochlorophyll and bacteriopheophytin in active A-branch respectively). The excited states emit at 905 and 940nm and have approximately the same energy and high interaction rate. The intermediate state I is populated earlier than the P+BA−HA state and has energy close to the energy of the excited states, a high rate of population and depopulation and spectral identity to the BA−. A sum of the I and P+BA−HA populations fits the experimental kinetics of the BA− absorption band at 1020nm. The model explains an oscillatory phenomenon in the kinetics of the P⁎ stimulated emission and of the BA− absorption. In the schemes without the I state, accordance with the experiment is achieved at unreal parameter values or is not achieved at all. A qualitative agreement of the model with the experiment can be achieved at a wide range of parameter values. The nature of the states I and P⁎940BAHA is discussed in terms of partial charge separation between P and BA and inside P respectively.

Assessment of wastewater and recycled water quality: A comparison of lines of evidence from in vitro, in vivo and chemical analyses

We investigated water quality at an advanced water reclamation plant and three conventional wastewater treatment plants using an “ecotoxicity toolbox” consisting of three complementary analyses (chemical analysis, in vitro bioanalysis and in situ biological monitoring), with a focus on endocrine disruption. The in vitro bioassays were chosen to provide an appropriately wide coverage of biological effects relevant to managed aquifer recharge and environmental discharge of treated wastewater, and included bioassays for bacterial toxicity (Microtox), genotoxicity (umuC), photosynthesis inhibition (Max-I-PAM) and endocrine effects (E-SCREEN and AR-CALUX). Chemical analysis of hormones and pesticides using LCMSMS was performed in parallel to correlate standard analytical methods with the in vitro assessment. For two plants with surface water discharge into open drains, further field work was carried out to examine in situ effects using mosquitofish (Gambusia holbrooki) as a bioindicator species for possible endocrine effects. The results show considerable cytotoxicity, phytotoxicity, estrogenicity and androgenicity in raw sewage, all of which were significantly reduced by conventional wastewater treatment. No biological response was detected to RO water, suggesting that reverse osmosis is a significant barrier to biologically active compounds. Chemical analysis and in situ monitoring revealed trends consistent with the in vitro results: chemical analysis confirmed the removal trends observed by the bioanalytical tools, and in situ sampling did not reveal any evidence of endocrine disruption specifically due to discharge of treated wastewater (although other sources may be present). Biomarkers of exposure (in vitro) and effect (in vivo or in situ) are complementary and together provide information with a high level of ecological relevance. This study illustrates the utility of combining multiple lines of evidence in the assessment of water quality.

Three-Dimensional Structure of the Rhodobacter sphaeroides RC-LH1-PufX Complex: Dimerization and Quinone Channels Promoted by PufX

Reaction center-light harvesting 1 (RC-LH1) complexes are the fundamental units of bacterial photosynthesis, which use solar energy to power the reduction of quinone to quinol prior to the formation of the proton gradient that drives ATP synthesis. The dimeric RC-LH1-PufX complex of Rhodobacter sphaeroides is composed of 64 polypeptides and 128 cofactors, including 56 LH1 bacteriochlorophyll a (BChl a) molecules that surround and donate energy to the two RCs. The 3D structure was determined to 8 Å by X-ray crystallography, and a model was built with constraints provided by electron microscopy (EM), nuclear magnetic resonance (NMR), mass spectrometry (MS), and site-directed mutagenesis. Each half of the dimer complex consists of a RC surrounded by an array of 14 LH1 αβ subunits, with two BChls sandwiched between each αβ pair of transmembrane helices. The N- and C-terminal extrinsic domains of PufX promote dimerization by interacting with the corresponding domains of an LH1 β polypeptide from the other half of the RC-LH1-PufX complex. Close contacts between PufX, an LH1 αβ subunit, and the cytoplasmic domain of the RC-H subunit prevent the LH1 complex from encircling the RC and create a channel connecting the RC QB site to an opening in the LH1 ring, allowing Q/QH₂ exchange with the external quinone pool. We also identified a channel that connects the two halves of the dimer, potentially forming a long-range pathway for quinone migration along rows of RC-LH1-PufX complexes in the membrane. The structure of the RC-LH1-PufX complex explains the crucial role played by PufX in dimer formation, and it shows how quinone traffic traverses the LH1 complex as it shuttles between the RC and the cytochrome bc₁ complex.

Whole-Genome Sequence of the Purple Photosynthetic Bacterium Rhodovulum sulfidophilum Strain W4

We report the draft genome sequence of the purple photosynthetic bacterium Rhodovulum sulfidophilum. The photosynthesis gene cluster comprises two segments—a unique feature among photosynthesis gene clusters of purple bacteria. The genome information will be useful for further analysis of bacterial photosynthesis.

The three-dimensional structures of bacterial reaction centers

This review presents a broad overview of the research that enabled the structure determination of the bacterial reaction centers from Blastochloris viridis and Rhodobacter sphaeroides, with a focus on the contributions from Duysens, Clayton, and Feher. Early experiments performed in the laboratory of Duysens and others demonstrated the utility of spectroscopic techniques and the presence of photosynthetic complexes in both oxygenic and anoxygenic photosynthesis. The laboratories of Clayton and Feher led efforts to isolate and characterize the bacterial reaction centers. The availability of well-characterized preparations of pure and stable reaction centers allowed the crystallization and subsequent determination of the structures using X-ray diffraction. The three-dimensional structures of reaction centers revealed an overall arrangement of two symmetrical branches of cofactors surrounded by transmembrane helices from the L and M subunits, which also are related by the same twofold symmetry axis. The structure has served as a framework to address several issues concerning bacterial photosynthesis, including the directionality of electron transfer, the properties of the reaction center-cytochrome c 2 complex, and the coupling of proton and electron transfer. Together, these research efforts laid the foundation for ongoing efforts to address an outstanding question in oxygenic photosynthesis, namely the molecular mechanism of water oxidation.

Photophysical properties and electronic structure of bacteriochlorin-chalcones with extended near-infrared absorption.

Synthetic bacteriochlorins enable systematic tailoring of substituents about the bacteriochlorin chromophore and thereby provide insights concerning the native bacteriochlorophylls of bacterial photosynthesis. Nine free-base bacteriochlorins (eight prepared previously and one prepared here) have been examined that bear diverse substituents at the 13- or 3,13-positions. The substituents include chalcone (3-phenylprop-2-en-1-onyl) derivatives with groups attached to the phenyl moiety, a "reverse chalcone" (3-phenyl-3-oxo-1-enyl), and extended chalcones (5-phenylpenta-2,4-dien-1-onyl, retinylidenonyl). The spectral and photophysical properties (τs, Φf, Φ(ic), Φ(isc), τT, k(f), k(ic), k(isc)) of the bacteriochlorins have been characterized. The bacteriochlorins absorb strongly in the 780-800 nm region and have fluorescence quantum yields (Φf) in the range 0.05-0.11 in toluene and dimethylsulfoxide. Light-induced electron promotions between orbitals with predominantly substituent or macrocycle character or both may give rise to some net macrocycle ↔ substituent charge-transfer character in the lowest and higher singlet excited states as indicated by density functional theory (DFT) and time-dependent DFT calculations. Such calculations indicated significant participation of molecular orbitals beyond those (HOMO - 1 to LUMO + 1) in the Gouterman four-orbital model. Taken together, the studies provide insight into the fundamental properties of bacteriochlorins and illustrate designs for tuning the spectral and photophysical features of these near-infrared-absorbing tetrapyrrole chromophores.