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1.
Nat Commun ; 11(1): 1542, 2020 03 24.
Artigo em Inglês | MEDLINE | ID: mdl-32210238

RESUMO

Natural photosynthesis can be divided between the chlorophyll-containing plants, algae and cyanobacteria that make up the oxygenic phototrophs and a diversity of bacteriochlorophyll-containing bacteria that make up the anoxygenic phototrophs. Photosynthetic light harvesting and reaction centre proteins from both kingdoms have been exploited for solar energy conversion, solar fuel synthesis and sensing technologies, but the energy harvesting abilities of these devices are limited by each protein's individual palette of pigments. In this work we demonstrate a range of genetically-encoded, self-assembling photosystems in which recombinant plant light harvesting complexes are covalently locked with reaction centres from a purple photosynthetic bacterium, producing macromolecular chimeras that display mechanisms of polychromatic solar energy harvesting and conversion. Our findings illustrate the power of a synthetic biology approach in which bottom-up construction of photosystems using naturally diverse but mechanistically complementary components can be achieved in a predictable fashion through the encoding of adaptable, plug-and-play covalent interfaces.


Assuntos
Proteínas de Arabidopsis/química , Proteínas de Bactérias/química , Bacterioclorofilas/química , Complexos de Proteínas Captadores de Luz/química , Energia Solar , Biologia Sintética/métodos , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/efeitos da radiação , Proteínas de Bactérias/genética , Proteínas de Bactérias/efeitos da radiação , Bacterioclorofilas/genética , Bacterioclorofilas/efeitos da radiação , Carotenoides/química , Carotenoides/efeitos da radiação , Complexos de Proteínas Captadores de Luz/genética , Complexos de Proteínas Captadores de Luz/efeitos da radiação , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/efeitos da radiação , Rhodobacter sphaeroides/química , Rhodobacter sphaeroides/genética , Rhodobacter sphaeroides/efeitos da radiação , Luz Solar
2.
Nucleic Acids Res ; 48(6): e33, 2020 04 06.
Artigo em Inglês | MEDLINE | ID: mdl-31989175

RESUMO

Light-regulated modules offer unprecedented new ways to control cellular behaviour with precise spatial and temporal resolution. Among a variety of bacterial light-switchable gene expression systems, single-component systems consisting of single transcription factors would be more useful due to the advantages of speed, simplicity, and versatility. In the present study, we developed a single-component light-activated bacterial gene expression system (eLightOn) based on a novel LOV domain from Rhodobacter sphaeroides (RsLOV). The eLightOn system showed significant improvements over the existing single-component bacterial light-activated expression systems, with benefits including a high ON/OFF ratio of >500-fold, a high activation level, fast activation kinetics, and/or good adaptability. Additionally, the induction characteristics, including regulatory windows, activation kinetics and light sensitivities, were highly tunable by altering the expression level of LexRO. We demonstrated the usefulness of the eLightOn system in regulating cell division and swimming by controlling the expression of the FtsZ and CheZ genes, respectively, as well as constructing synthetic Boolean logic gates using light and arabinose as the two inputs. Taken together, our data indicate that the eLightOn system is a robust and highly tunable tool for quantitative and spatiotemporal control of bacterial gene expression.


Assuntos
Regulação Bacteriana da Expressão Gênica/efeitos da radiação , Luz , Rhodobacter sphaeroides/citologia , Rhodobacter sphaeroides/efeitos da radiação , Proteínas de Bactérias/metabolismo , Divisão Celular/efeitos da radiação , Cinética , Lógica , Fatores de Transcrição/metabolismo
3.
J Agric Food Chem ; 67(34): 9560-9568, 2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31368704

RESUMO

ß-Carotene is a precursor of vitamin A and a dietary supplement for its antioxidant property. Producing ß-carotene by microbial fermentation has attracted much attention owing to consumers' preference for the natural product. In this study, an engineered photosynthetic Rhodobacter sphaeroides producing ß-carotene was constructed by the following strategies: (1) five promoters of different strengths were used to investigate the effect of the expression level of crtY on ß-carotene content. It was found that PrrnB increased the ß-carotene content by 109%. (2) blocking of the branched pentose phosphate pathway by zwf deletion, and (3) overexpressing dxs could restore the transcriptional levels of crtE and crtB. Finally, the engineered RS-C3 has the highest ß-carotene content of 14.93 mg/g dry cell weight (DCW) among all of the reported photosynthetic bacteria and the ß-carotene content reached 3.34 mg/g DCW under light conditions. Our results will be available for industrial use to supply a large quantity of natural ß-carotene.


Assuntos
Proteínas de Bactérias/genética , Liases Intramoleculares/genética , Rhodobacter sphaeroides/genética , Rhodobacter sphaeroides/metabolismo , beta Caroteno/biossíntese , Proteínas de Bactérias/metabolismo , Fermentação , Liases Intramoleculares/metabolismo , Luz , Engenharia Metabólica , Regiões Promotoras Genéticas , Rhodobacter sphaeroides/efeitos da radiação
4.
Enzyme Microb Technol ; 110: 1-7, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29310850

RESUMO

In this study, distillery wastewater was treated by dark fermentation or photofermentation alone, and by sequential dark and photofermentation processes using anaerobic saccharolytic consortium and purple nonsulfur bacteria. Combination of dark and photofermentation resulted in the maximal H2 yield of 17.6L/L of distillery waste with chemical oxygen demand 40g/L. It is equivalent to 205kJ/L distillery wastewater and corresponds to recovery of approximately 4-8% of energy consumed during ethanol production. Optimal performance of photofermentation was observed at 20% concentration of pre-fermented distillery waste. In photofermentation, the range of the suitable distillery waste concentrations was extended and the H2 yield was improved by choosing the tolerant strain of purple bacteria Rhodobacter sphaeroides B-3059. After two stages, organic acids and sugars were completely consumed that means wastewater treatment concomitant to H2 production.


Assuntos
Fermentação , Hidrogênio/metabolismo , Rhodobacter capsulatus/metabolismo , Rhodobacter sphaeroides/metabolismo , Águas Residuárias/microbiologia , Concentração de Íons de Hidrogênio , Luz , Rhodobacter capsulatus/crescimento & desenvolvimento , Rhodobacter capsulatus/efeitos da radiação , Rhodobacter sphaeroides/crescimento & desenvolvimento , Rhodobacter sphaeroides/efeitos da radiação , Águas Residuárias/química
5.
Photosynth Res ; 136(3): 379-392, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29285578

RESUMO

Mercuric contamination of aqueous cultures results in impairment of viability of photosynthetic bacteria primarily by inhibition of the photochemistry of the reaction center (RC) protein. Isolated reaction centers (RCs) from Rhodobacter sphaeroides were exposed to Hg2+ ions up to saturation concentration (~ 103 [Hg2+]/[RC]) and the gradual time- and concentration-dependent loss of the photochemical activity was monitored. The vast majority of Hg2+ ions (about 500 [Hg2+]/[RC]) had low affinity for the RC [binding constant Kb ~ 5 mM-1] and only a few (~ 1 [Hg2+]/[RC]) exhibited strong binding (Kb ~ 50 µM-1). Neither type of binding site had specific and harmful effects on the photochemistry of the RC. The primary charge separation was preserved even at saturation mercury(II) concentration, but essential further steps of stabilization and utilization were blocked already in the 5 < [Hg2+]/[RC] < 50 range whose locations were revealed. (1) The proton gate at the cytoplasmic site had the highest affinity for Hg2+ binding (Kb ~ 0.2 µM-1) and blocked the proton uptake. (2) Reduced affinity (Kb ~ 0.05 µM-1) was measured for the mercury(II)-binding site close to the secondary quinone that resulted in inhibition of the interquinone electron transfer. (3) A similar affinity was observed close to the bacteriochlorophyll dimer causing slight energetic changes as evidenced by a ~ 30 nm blue shift of the red absorption band, a 47 meV increase in the redox midpoint potential, and a ~ 20 meV drop in free energy gap of the primary charge pair. The primary quinone was not perturbed upon mercury(II) treatment. Although the Hg2+ ions attack the RC in large number, the exertion of the harmful effect on photochemistry is not through mass action but rather a couple of well-defined targets. Bound to these sites, the Hg2+ ions can destroy H-bond structures, inhibit protein dynamics, block conformational gating mechanisms, and modify electrostatic profiles essential for electron and proton transfer.


Assuntos
Transporte de Elétrons/efeitos da radiação , Mercúrio/farmacologia , Complexo de Proteínas do Centro de Reação Fotossintética/efeitos dos fármacos , Complexo de Proteínas do Centro de Reação Fotossintética/efeitos da radiação , Prótons , Rhodobacter sphaeroides/efeitos dos fármacos , Bacterioclorofilas/metabolismo , Benzoquinonas/metabolismo , Sítios de Ligação , Fotoquímica , Fotossíntese/efeitos dos fármacos , Rhodobacter sphaeroides/fisiologia , Rhodobacter sphaeroides/efeitos da radiação , Água/metabolismo
6.
Nat Commun ; 8(1): 988, 2017 10 17.
Artigo em Inglês | MEDLINE | ID: mdl-29042567

RESUMO

Photosynthesis transfers energy efficiently through a series of antenna complexes to the reaction center where charge separation occurs. Energy transfer in vivo is primarily monitored by measuring fluorescence signals from the small fraction of excitations that fail to result in charge separation. Here, we use two-dimensional electronic spectroscopy to follow the entire energy transfer process in a thriving culture of the purple bacteria, Rhodobacter sphaeroides. By removing contributions from scattered light, we extract the dynamics of energy transfer through the dense network of antenna complexes and into the reaction center. Simulations demonstrate that these dynamics constrain the membrane organization into small pools of core antenna complexes that rapidly trap energy absorbed by surrounding peripheral antenna complexes. The rapid trapping and limited back transfer of these excitations lead to transfer efficiencies of 83% and a small functional light-harvesting unit.During photosynthesis, energy is transferred from photosynthetic antenna to reaction centers via ultrafast energy transfer. Here the authors track energy transfer in photosynthetic bacteria using two-dimensional electronic spectroscopy and show that these transfer dynamics constrain antenna complex organization.


Assuntos
Transferência de Energia , Fotossíntese/fisiologia , Rhodobacter sphaeroides/metabolismo , Energia Solar , Proteínas de Bactérias/metabolismo , Fluorescência , Cinética , Luz , Complexo de Proteínas do Centro de Reação Fotossintética/metabolismo , Proteobactérias/citologia , Proteobactérias/metabolismo , Proteobactérias/efeitos da radiação , Rhodobacter sphaeroides/citologia , Rhodobacter sphaeroides/efeitos da radiação , Espectrofotometria/métodos
7.
Biochemistry (Mosc) ; 82(8): 906-915, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28941458

RESUMO

Energy relaxation was studied with difference femtosecond spectroscopy in reaction centers of the YM210L mutant of the purple photosynthetic bacterium Rhodobacter sphaeroides at low temperature (90 K). A dynamical long-wavelength shift of stimulated emission of the excited state of the bacteriochlorophyll dimer P was found, which starts simultaneously with P* formation and is accompanied by a change in the spectral shape of this emission. The characteristic value of this shift was about 30 nm, and the characteristic time about 200 fs. Difference kinetics ΔA measured at fixed wavelengths demonstrate the femtosecond shift of the P* stimulated emission appearing as a dependence of these kinetics on wavelength. We found that the reported long-wavelength shift can be explained in terms of electron-vibrational relaxation of the P* excited state with time constants of vibrational and electronic relaxation of 100 and 50 fs, respectively. Alternative mechanisms of the dynamical shift of the P* stimulated emission spectrum are also discussed in terms of energy redistribution between vibrational modes or coherent excitation of the modes.


Assuntos
Proteínas de Bactérias/metabolismo , Complexo de Proteínas do Centro de Reação Fotossintética/metabolismo , Rhodobacter sphaeroides/metabolismo , Proteínas de Bactérias/genética , Bacterioclorofilas/química , Bacterioclorofilas/metabolismo , Dimerização , Cinética , Lasers de Estado Sólido , Mutagênese Sítio-Dirigida , Complexo de Proteínas do Centro de Reação Fotossintética/genética , Rhodobacter sphaeroides/efeitos da radiação , Espectrofotometria
8.
Biochim Biophys Acta Bioenerg ; 1858(9): 795-803, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-28587931

RESUMO

In bacterial photosynthesis reaction center-light-harvesting 1 (RC-LH1) complexes trap absorbed solar energy by generating a charge separated state. Subsequent electron and proton transfers form a quinol, destined to diffuse to the cytochrome bc1 complex. In bacteria such as Rhodobacter (Rba.) sphaeroides and Rba. capsulatus the PufX polypeptide creates a channel for quinone/quinol traffic across the LH1 complex that surrounds the RC, and it is therefore essential for photosynthetic growth. PufX also plays a key role in dimerization of the RC-LH1-PufX core complex, and the structure of the Rba. sphaeroides complex shows that the PufX C-terminus, particularly the region from X49-X53, likely mediates association of core monomers. To investigate this putative interaction we analysed mutations PufX R49L, PufX R53L, PufX R49/53L and PufX G52L by measuring photosynthetic growth, fractionation of detergent-solubilised membranes, formation of 2-D crystals and electron microscopy. We show that these mutations do not affect assembly of PufX within the core or photosynthetic growth but they do prevent dimerization, consistent with predictions from the RC-LH1-PufX structure. We obtained low resolution structures of monomeric core complexes with and without PufX, using electron microscopy of negatively stained single particles and 3D reconstruction; the monomeric complex with PufX corresponds to one half of the dimer structure whereas LH1 completely encloses the RC if the gene encoding PufX is deleted. On the basis of the insights gained from these mutagenesis and structural analyses we propose a sequence for assembly of the dimeric RC-LH1-PufX complex.


Assuntos
Proteínas de Bactérias/fisiologia , Complexos de Proteínas Captadores de Luz/química , Rhodobacter sphaeroides/metabolismo , Sequência de Aminoácidos , Substituição de Aminoácidos , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/ultraestrutura , Benzoquinonas/metabolismo , Cristalização , Dimerização , Hidroquinonas/metabolismo , Processamento de Imagem Assistida por Computador , Complexos de Proteínas Captadores de Luz/genética , Complexos de Proteínas Captadores de Luz/fisiologia , Complexos de Proteínas Captadores de Luz/ultraestrutura , Microscopia Eletrônica , Modelos Moleculares , Mutação de Sentido Incorreto , Mutação Puntual , Conformação Proteica , Domínios Proteicos , Rhodobacter sphaeroides/genética , Rhodobacter sphaeroides/efeitos da radiação
9.
Dokl Biochem Biophys ; 473(1): 118-121, 2017 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-28510131

RESUMO

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


Assuntos
Dipiridamol/metabolismo , Luz , Complexo de Proteínas do Centro de Reação Fotossintética/metabolismo , Rhodobacter sphaeroides/metabolismo , Rhodobacter sphaeroides/efeitos da radiação , Radicais Livres/metabolismo , Cinética , Rhodobacter sphaeroides/enzimologia
10.
Photosynth Res ; 133(1-3): 371-377, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-28540587

RESUMO

Bacterial reaction centers (RC) from Rhodobacter sphaeroides have been widely used to functionalize electrodes and to generate photocurrent. However, in most studies, direct electron transfer from the semiquinone to the electrode was not observed because the H subunit of the RC shields the semiquinone. It is demonstrated in the current work that removal of the H subunit effectively exposes the semiquinone sites in the LM dimer. This is demonstrated by measuring the second-order rate constant for the reaction between ferricyanide and the anionic semiquinone Q A- formed by an actinic flash. The rate constant increases 1000-fold for Q A- oxidation by ferricyanide in the LM dimer compared to the intact RC. The second-order rate constant approaches the diffusion limit of 6 × 109 M-1·s-1 at low pH, but it decreases steadily when the pH is above 6.5. This pH dependence suggests that the protonation state of the LM dimer plays an important role in controlling the electron transfer kinetics. It is also shown that the addition of exogenous ubiquinone to replenish the QB site, which is mostly empty in the LM dimer, leads to oxidation of Q A- by O2 following an actinic flash. It is concluded that removal of the H subunit results in exposure of the semiquinone sites of the LM dimer to externally added oxidants and may provide a strategy for enhancing direct electron transfer from the RC to an electrode.


Assuntos
Benzoquinonas/metabolismo , Ferricianetos/farmacologia , Oxigênio/farmacologia , Multimerização Proteica , Subunidades Proteicas/metabolismo , Rhodobacter sphaeroides/metabolismo , Transporte de Elétrons/efeitos da radiação , Elétrons , Concentração de Íons de Hidrogênio , Cinética , Luz , Oxirredução , Rhodobacter sphaeroides/efeitos da radiação , Cloreto de Sódio/química
11.
Proc Natl Acad Sci U S A ; 114(7): 1480-1485, 2017 02 14.
Artigo em Inglês | MEDLINE | ID: mdl-28137837

RESUMO

Blue light using flavin adenine dinucleotide (BLUF) proteins are essential for the light regulation of a variety of physiologically important processes and serve as a prototype for photoinduced proton-coupled electron transfer (PCET). Free-energy simulations elucidate the active site conformations in the AppA (activation of photopigment and puc expression) BLUF domain before and following photoexcitation. The free-energy profile for interconversion between conformations with either Trp104 or Met106 closer to the flavin, denoted Trpin/Metout and Trpout/Metin, reveals that both conformations are sampled on the ground state, with the former thermodynamically favorable by ∼3 kcal/mol. These results are consistent with the experimental observation of both conformations. To analyze the proton relay from Tyr21 to the flavin via Gln63, the free-energy profiles for Gln63 rotation were calculated on the ground state, the locally excited state of the flavin, and the charge-transfer state associated with electron transfer from Tyr21 to the flavin. For the Trpin/Metout conformation, the hydrogen-bonding pattern conducive to the proton relay is not thermodynamically favorable on the ground state but becomes more favorable, corresponding to approximately half of the configurations sampled, on the locally excited state. The calculated energy gaps between the locally excited and charge-transfer states suggest that electron transfer from Tyr21 to the flavin is more facile for configurations conducive to proton transfer. When the active site conformation is not conducive to PCET from Tyr21, Trp104 can directly compete with Tyr21 for electron transfer to the flavin through a nonproductive pathway, impeding the signaling efficiency.


Assuntos
Proteínas de Bactérias/química , Simulação por Computador , Flavoproteínas/química , Fotorreceptores Microbianos/química , Rhodobacter sphaeroides/metabolismo , Proteínas de Bactérias/efeitos da radiação , Domínio Catalítico , Transporte de Elétrons , Mononucleotídeo de Flavina/química , Flavoproteínas/efeitos da radiação , Glutamina/química , Ligação de Hidrogênio , Luz , Metionina/química , Modelos Moleculares , Fotorreceptores Microbianos/efeitos da radiação , Conformação Proteica/efeitos da radiação , Domínios Proteicos , Rhodobacter sphaeroides/efeitos da radiação , Triptofano/química , Tirosina/química , Tirosina/efeitos da radiação
12.
FEBS Lett ; 591(4): 573-580, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-28130884

RESUMO

Photosynthesis in some phototrophic bacteria requires the PufX component of the reaction centre-light-harvesting 1-PufX (RC-LH1-PufX) complex, which creates a pore for quinone/quinol (Q/QH2 ) exchange across the LH1 barrier surrounding the RC. However, photosynthetic bacteria such as Thermochromatium (T.) tepidum do not require PufX because there are fewer carotenoid binding sites, which creates multiple pores in the LH1 ring for Q/QH2 exchange. We show that an αTrp-24 →Phe alteration of the Rhodobacter (Rba.) sphaeroides LH1 antenna impairs carotenoid binding and allows photosynthetic growth in the absence of PufX. We propose that acquisition of PufX and confining Q/QH2 traffic to a pore adjacent to the RC QB site is an evolutionary upgrade that allows increased LH1 carotenoid content for enhanced light absorption and photoprotection.


Assuntos
Proteínas de Bactérias/metabolismo , Benzoquinonas/metabolismo , Carotenoides/metabolismo , Complexos de Proteínas Captadores de Luz/metabolismo , Rhodobacter sphaeroides/metabolismo , Proteínas de Bactérias/genética , Bacterioclorofilas/metabolismo , Luz , Complexos de Proteínas Captadores de Luz/genética , Mutação , Fotossíntese/genética , Fotossíntese/efeitos da radiação , Ligação Proteica , Rhodobacter sphaeroides/genética , Rhodobacter sphaeroides/efeitos da radiação , Espectrofotometria
13.
Biochim Biophys Acta ; 1857(12): 1925-1934, 2016 12.
Artigo em Inglês | MEDLINE | ID: mdl-27687473

RESUMO

Upon photoexcitation, the reaction center (RC) pigment-proteins that facilitate natural photosynthesis achieve a metastable separation of electrical charge among the embedded cofactors. Because of the high quantum efficiency of this process, there is a growing interest in their incorporation into biohybrid materials for solar energy conversion, bioelectronics and biosensing. Multiple bioelectrochemical studies have shown that reaction centers from various photosynthetic organisms can be interfaced with diverse electrode materials for the generation of photocurrents, but many mechanistic aspects of native protein functionality in a non-native environment is unknown. In vivo, RC's catalyse ubiquinone-10 reduction, protonation and exchange with other lipid phase ubiquinone-10s via protein-controlled spatial orientation and protein rearrangement. In contrast, the mechanism of ubiquinone-0 reduction, used to facilitate fast RC turnover in an aqueous photoelectrochemical cell (PEC), may not proceed via the same pathway as the native cofactor. In this report we show truncation of the native isoprene tail results in larger RC turnover rates in a PEC despite the removal of the tail's purported role of ubiquinone headgroup orientation and binding. Through the use of reaction centers with single or double mutations, we also show the extent to which two-electron/two-proton ubiquinone chemistry that operates in vivo also underpins the ubiquinone-0 reduction by surface-adsorbed RCs in a PEC. This reveals that only the ubiquinone headgroup is critical to the fast turnover of the RC in a PEC and provides insight into design principles for the development of new biophotovoltaic cells and biosensors.


Assuntos
Eletroquímica/métodos , Luz , Fotossíntese/efeitos da radiação , Complexo de Proteínas do Centro de Reação Fotossintética/efeitos da radiação , Rhodobacter sphaeroides/efeitos da radiação , Ubiquinona/efeitos da radiação , Técnicas Biossensoriais , Eletroquímica/instrumentação , Eletrodos , Transporte de Elétrons , Cinética , Modelos Biológicos , Mutação , Oxirredução , Complexo de Proteínas do Centro de Reação Fotossintética/química , Complexo de Proteínas do Centro de Reação Fotossintética/genética , Complexo de Proteínas do Centro de Reação Fotossintética/metabolismo , Conformação Proteica , Rhodobacter sphaeroides/genética , Rhodobacter sphaeroides/metabolismo , Energia Solar , Relação Estrutura-Atividade , Ubiquinona/metabolismo
14.
Elife ; 52016 08 26.
Artigo em Inglês | MEDLINE | ID: mdl-27564854

RESUMO

The chromatophore of purple bacteria is an intracellular spherical vesicle that exists in numerous copies in the cell and that efficiently converts sunlight into ATP synthesis, operating typically under low light conditions. Building on an atomic-level structural model of a low-light-adapted chromatophore vesicle from Rhodobacter sphaeroides, we investigate the cooperation between more than a hundred protein complexes in the vesicle. The steady-state ATP production rate as a function of incident light intensity is determined after identifying quinol turnover at the cytochrome bc1 complex (cytb⁢c1) as rate limiting and assuming that the quinone/quinol pool of about 900 molecules acts in a quasi-stationary state. For an illumination condition equivalent to 1% of full sunlight, the vesicle exhibits an ATP production rate of 82 ATP molecules/s. The energy conversion efficiency of ATP synthesis at illuminations corresponding to 1%-5% of full sunlight is calculated to be 0.12-0.04, respectively. The vesicle stoichiometry, evolutionarily adapted to the low light intensities in the habitat of purple bacteria, is suboptimal for steady-state ATP turnover for the benefit of protection against over-illumination.


Assuntos
Trifosfato de Adenosina/biossíntese , Cromatóforos Bacterianos/metabolismo , Cromatóforos Bacterianos/efeitos da radiação , Metabolismo Energético , Rhodobacter sphaeroides/metabolismo , Rhodobacter sphaeroides/efeitos da radiação , Hidroquinonas/análise , Luz , Quinonas/análise
15.
J Photochem Photobiol B ; 162: 592-596, 2016 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-27479839

RESUMO

The present work was focused on the effects of low-intensity (the flux capacity was of 0.06mWcm(-2)) electromagnetic irradiation (EMI) of extremely high frequencies or millimeter waves on the growth and hydrogen (H2) photoproduction by purple non-sulfur bacteria Rhodobacter sphaeroides MDC6521 (from Armenian mineral springs). After exposure of R. sphaeroides, grown under anaerobic conditions upon illumination, to EMI (51.8GHz and 53.0GHz) for 15min an increase of specific growth rate by ~1.2-fold, in comparison with control (non-irradiated cells), was obtained. However, the effect of EMI depends on the duration of irradiation: the exposure elongation up to 60min caused the delay of the growth lag phase and the decrease specific growth rate by ~1.3-fold, indicating the bactericidal effect of EMI. H2 yield of the culture, irradiated by EMI for 15min, determined during 72h growth, was ~1.2-fold higher than H2 yield of control cells, whereas H2 production by cultures, irradiated by EMI for 60min was not observed during 72h growth. This difference in the effects of extremely high frequency EMI indicates a direct effect of radiation on the membrane transfer and the enzymes of these bacteria. Moreover, EMI increased DCCD-inhibited H(+) fluxes across the bacterial membrane and DCCD-sensitive ATPase activity of membrane vesicles, indicating that the proton FoF1-ATPase is presumably a basic target for extremely high frequency EMI related to H2 production by cultures.


Assuntos
Radiação Eletromagnética , Hidrogênio/metabolismo , Rhodobacter sphaeroides/metabolismo , Proteínas de Bactérias/metabolismo , Parede Celular/metabolismo , Oxirredução , ATPases Translocadoras de Prótons/metabolismo , Rhodobacter sphaeroides/crescimento & desenvolvimento , Rhodobacter sphaeroides/efeitos da radiação
16.
Chemosphere ; 159: 138-144, 2016 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-27285383

RESUMO

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 (14)C-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.


Assuntos
Substâncias Explosivas/metabolismo , Luz , Rhodobacter sphaeroides/metabolismo , Rhodobacter sphaeroides/efeitos da radiação , Triazinas/metabolismo , Poluentes Químicos da Água/metabolismo , Antraquinonas/farmacologia , Biodegradação Ambiental/efeitos dos fármacos , Biodegradação Ambiental/efeitos da radiação , Cinética , Malatos/farmacologia , Oxirredução , Rhodobacter sphaeroides/efeitos dos fármacos , Ácido Succínico/farmacologia
17.
Dokl Biochem Biophys ; 467(1): 105-9, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-27193710

RESUMO

The differences in the average fluorescence lifetime (τav) of tryptophanyls in photosynthetic reaction center (RC) of the purple bacteria Rb. sphaeroides frozen to 80 K in the dark or on the actinic light was found. This difference disappeared during subsequent heating at the temperatures above 250 K. The computer-based calculation of vibration spectra of the tryptophan molecule was performed. As a result, the normal vibrational modes associated with deformational vibrations of the aromatic ring of the tryptophan molecule were found. These deformational vibrations may be active during the nonradiative transition of the molecule from the excited to the ground state. We assume that the differences in τav may be associated with the change in the activity of these vibration modes due to local variations in the microenvironment of tryptophanyls during the light activation.


Assuntos
Proteínas de Bactérias/metabolismo , Fluorescência , Complexo de Proteínas do Centro de Reação Fotossintética/metabolismo , Rhodobacter sphaeroides/metabolismo , Temperatura , Proteínas de Bactérias/química , Proteínas de Bactérias/efeitos da radiação , Glicerol/química , Modelos Moleculares , Complexo de Proteínas do Centro de Reação Fotossintética/química , Complexo de Proteínas do Centro de Reação Fotossintética/efeitos da radiação , Conformação Proteica , Rhodobacter sphaeroides/química , Rhodobacter sphaeroides/efeitos da radiação , Triptofano/química , Vibração , Água/química
18.
Photosynth Res ; 130(1-3): 307-316, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27034065

RESUMO

The composition of photosynthetic apparatus of Rhodobacter sphaeroides wild strain 2.4.1 and its LHII-deficient mutant DBCΩ was compared. The absence of LHII in the mutant was confirmed by comparison of chromatophores spectra and by the absence of electrophoretic band corresponding to LHII complex. Continuous turbidostat cultures of wild strain and its LHII-deficient mutant were compared in response to different light intensities. Cultures were grown using lactate, mixture of lactate and acetate or succinate as carbon source. For comparative analysis, an approximation of experimental data by Monod and Gompertz equations were used. Cultures of DBCΩ had lower growth rates than wild strain when grown on lactate as electron donor and carbon source. Cultures of both strains grown on lactate and acetate or on succinate had similar growth rates. The cultures showed maximum growth rates when grown with succinate. Bacteriochlorophyll a content increased in both strains with decrease of incident light intensity. However, the variation of Bchl a content in wild strain was much more significant. Under light-limiting conditions, bacteriochlorophyll a content in DBCΩ was 4-5 times lower than in the wild strain. Under light-saturating conditions, it was only 1.5-2.5 times lower. Growing with lactate or with lactate and acetate, the mutant switched from light limitation under low light intensities to limitation by organic acids under higher light, whereas the parental strain had similar switch of limiting factor only when growing with lactate and acetate mixture. DBCΩ mutant has higher minimal light intensity enabling growth on any organic acid as a substrate. When growing with lactate or with lactate and acetate, the mutant reached maximum growth rate at lower light intensities than the wild strain. This phenomenon was observed for the first time. Taking into account the concentration of BChl a under light-limiting conditions, the thickness of the suspension capable of effective light absorption could be increased by 4-5 times, which is favorable for intensive cultivation.


Assuntos
Rhodobacter sphaeroides/efeitos da radiação , Acetatos/metabolismo , Proteínas de Bactérias/fisiologia , Bacterioclorofila A/metabolismo , Ácido Láctico/metabolismo , Luz , Complexos de Proteínas Captadores de Luz/deficiência , Complexos de Proteínas Captadores de Luz/metabolismo , Complexos de Proteínas Captadores de Luz/fisiologia , Complexo de Proteína do Fotossistema II/fisiologia , Rhodobacter sphaeroides/efeitos dos fármacos , Rhodobacter sphaeroides/crescimento & desenvolvimento , Succinatos/metabolismo
19.
Biochim Biophys Acta ; 1857(6): 634-42, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-27013332

RESUMO

In the purple phototrophic bacterium Rhodobacter sphaeroides, light harvesting LH2 complexes transfer absorbed solar energy to RC-LH1-PufX core complexes, which are mainly found in the dimeric state. Many other purple phototrophs have monomeric core complexes and the basis for requiring dimeric cores is not fully established, so we analysed strains of Rba. sphaeroides that contain either native dimeric core complexes or altered monomeric cores harbouring a deletion of the first 12 residues from the N-terminus of PufX, which retains the PufX polypeptide but removes the major determinant of core complex dimerization. Membranes were purified from strains with dimeric or monomeric cores, and with either high or low levels of the LH2 complex. Samples were interrogated with absorption, steady-state fluorescence, and picosecond time-resolved fluorescence kinetic spectroscopies to reveal their light-harvesting and energy trapping properties. We find that under saturating excitation light intensity the photosynthetic membranes containing LH2 and monomeric core complexes have fluorescence lifetimes nearly twice that of membranes with LH2 plus dimeric core complexes. This trend of increased lifetime is maintained with RCs in the open state as well, and for two different levels of LH2 content. Thus, energy trapping is more efficient when photosynthetic membranes of Rba. sphaeroides consist of RC-LH1-PufX dimers and LH2 complexes.


Assuntos
Cromatóforos Bacterianos/metabolismo , Proteínas de Bactérias/metabolismo , Complexos de Proteínas Captadores de Luz/metabolismo , Rhodobacter sphaeroides/metabolismo , Algoritmos , Cromatóforos Bacterianos/efeitos da radiação , Proteínas de Bactérias/química , Transferência de Energia/efeitos da radiação , Cinética , Luz , Complexos de Proteínas Captadores de Luz/química , Modelos Biológicos , Fotossíntese/efeitos da radiação , Multimerização Proteica/efeitos da radiação , Rhodobacter sphaeroides/efeitos da radiação , Espectrofotometria
20.
Enzyme Microb Technol ; 86: 45-51, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-26992792

RESUMO

The purple bacteria Rhodobacter sphaeroides serve as a promising biocatalyst in the photo-microbial fuel cell system (photo-MFC). This gram-negative species performs highly efficient anoxygenic photosynthesis that ensures an anaerobic environment in the anode compartment. Previous studies incorporating R. sphaeroides into photo-MFC were conducted using platinum as the anode electrode. In this study, we detected a steady current generation of R. sphaeroides in a bioelectrochemical system where glassy carbon was the working electrode and a typical growth medium was the electrolyte. The bioelectricity generation synchronized with the supplementation of reduced carbon source and showed immediate response to illumination, which strongly indicated the correlation between the observed current and the cytoplasmic quinone activity. Modifications of the endogenous electron flows mediated by quinone pool are shown to have significantly enhanced the bioelectricity generation. We anticipate that the findings in this study would advance future optimization of R. sphaeroides as an anode strain, as well as facilitate the study of bioenergetics in photosynthetic bacteria.


Assuntos
Fontes de Energia Bioelétrica/microbiologia , Rhodobacter sphaeroides/metabolismo , Biocatálise , Eletricidade , Transporte de Elétrons , Modelos Biológicos , Mutação , Fotossíntese , Quinonas/metabolismo , Rhodobacter sphaeroides/genética , Rhodobacter sphaeroides/efeitos da radiação , Ácido Succínico/metabolismo
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