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Emma Barahona, Emilio Jiménez-Vicente, Luis M Rubio
When produced biologically, especially by photosynthetic organisms, hydrogen gas (H2) is arguably the cleanest fuel available. An important limitation to the discovery or synthesis of better H2-producing enzymes is the absence of methods for the high-throughput screening of H2 production in biological systems. Here, we re-engineered the natural H2 sensing system of Rhodobacter capsulatus to direct the emission of LacZ-dependent fluorescence in response to nitrogenase-produced H2. A lacZ gene was placed under the control of the hupA H2-inducible promoter in a strain lacking the uptake hydrogenase and the nifH nitrogenase gene...
December 2, 2016: Scientific Reports
Emre M Ipekoğlu, Koray Göçmen, Mehmet T Öz, Muazzez Gürgan, Meral Yücel
Rhodobacter sphaeroides is a purple non-sulfur bacterium which photoheterotrophically produces hydrogen from organic acids under anaerobic conditions. A gene coding for putative chlorophyll a synthase (chlG) from cyanobacterium Prochlorococcus marinus was amplified by nested polymerase chain reaction and cloned into an inducible-expression plasmid which was subsequently transferred to R. sphaeroides for heterologous expression. Induced expression of chlG in R. sphaeroides led to changes in light absorption spectrum within 400-700 nm...
November 30, 2016: Journal of Basic Microbiology
Nadine Katzke, Andreas Knapp, Anita Loeschcke, Thomas Drepper, Karl-Erich Jaeger
Functional expression of genes from metagenomic libraries is limited by various factors including inefficient transcription and/or translation of target genes as well as improper folding and assembly of the corresponding proteins caused by the lack of appropriate chaperones and cofactors. It is now well accepted that the use of different expression hosts of distinct phylogeny and physiology can dramatically increase the rate of success. In the following chapter, we therefore describe tools and protocols allowing for the comparative heterologous expression of genes in five bacterial expression hosts, namely Escherichia coli, Pseudomonas putida, Bacillus subtilis, Burkholderia glumae, and Rhodobacter capsulatus...
2017: Methods in Molecular Biology
Ju-Yong Park, Yang-Hoon Kim, Jiho Min
In Rhodobacter sphaeroides, carbonic anhydrase (CA; EC is a zinc-containing metalloenzyme that catalyzes the reversible hydration of CO2 to HCO3(-) while phosphoenolpyruvate carboxylase (PEPC;, an enzyme involved in the carbon metabolism that catalyzed the fixation of CO2 to PEP, is a key factor for biological fixation of CO2 and enhances the production of organic compounds. In this study, the recombinant R. sphaeroides with highly-expressed CA was developed based on a surface displayed system of CA (pJY-OmpCA) on the outer membrane of R...
January 2017: Enzyme and Microbial Technology
Roberto Borghese, Marco Brucale, Gianuario Fortunato, Massimiliano Lanzi, Alessio Mezzi, Francesco Valle, Massimiliano Cavallini, Davide Zannoni
The toxic oxyanion tellurite (TeO3(2-)) is acquired by cells of Rhodobacter capsulatus grown anaerobically in the light, via acetate permease ActP2 and then reduced to Te(0) in the cytoplasm as needle-like black precipitates. Interestingly, photosynthetic cultures of R. capsulatus can also generate Te(0) nanoprecipitates (TeNPs) outside the cells upon addition of the redox mediator lawsone (2-hydroxy-1,4-naphtoquinone). TeNPs generation kinetics were monitored to define the optimal conditions to produce TeNPs as a function of various carbon sources and lawsone concentration...
November 15, 2016: Journal of Hazardous Materials
Kai Kudoh, Genma Kubota, Rintaro Fujii, Yusuke Kawano, Masaki Ihara
1-Deoxy-d-xylulose 5-phosphate synthase (DXS) is a rate-limiting enzyme in the 2-C-methyl-d-erythritol 4-phosphate (MEP) pathway, which is responsible for production of two precursors of all isoprenoids, isopentenyl diphosphate and dimethylallyl diphosphate (DMAPP). Previously, we attempted the overexpression of endogenous DXS in Synechocystis sp. PCC6803, and revealed that although the mRNA level was 4-fold higher, the DXS protein level was only 1.5-fold higher compared with those of the original strain, suggesting the lability of endogenous DXS protein...
November 14, 2016: Journal of Bioscience and Bioengineering
Dariusz M Niedzwiedzki, Preston L Dilbeck, Qun Tang, Elizabeth C Martin, David F Bocian, C Neil Hunter, Dewey Holten
Light-harvesting complex 2 (LH2) from the semi-aerobically grown purple phototrophic bacterium Rhodobacter sphaeroides was studied using optical (static and time-resolved) and resonance Raman spectroscopies. This antenna complex comprises bacteriochlorophyll (BChl) a and the carotenoid spheroidenone, a ketolated derivative of spheroidene. The results indicate that the spheroidenone-LH2 complex contains two spectral forms of the carotenoid: (1) a minor, "blue" form with an S2 (1(1)B u(+) ) spectral origin band at 522 nm, shifted from the position in organic media simply by the high polarizability of the binding site, and (2) the major, "red" form with the origin band at 562 nm that is associated with a pool of pigments that more strongly interact with protein residues, most likely via hydrogen bonding...
November 16, 2016: Photosynthesis Research
Julia Hahn, Sebastian Thalmann, Anzhela Migur, Raphael Freiherr von Boeselager, Nina Kubatova, Elena Kubareva, Harald Schwalbe, Elena Evguenieva-Hackenberg
Up to now, very small protein-coding genes have remained unrecognized in sequenced genomes. We identified an mRNA of 165 nucleotides (nt), which is conserved in Bradyrhizobiaceae and encodes a polypeptide with 14 amino acid residues (aa). The small mRNA harboring a unique Shine-Dalgarno sequence (SD) with a length of 17 nt was localized predominantly in the ribosome-containing P100 fraction of Bradyrhizobium japonicum USDA 110. Strong interaction between the mRNA and 30S ribosomal subunits was demonstrated by their co-sedimentation in sucrose density gradient...
November 11, 2016: RNA Biology
Folmer Fredslund, Harm Otten, Sabrina Gemperlein, Jens Christian N Poulsen, Yvonne Carius, Gert Wieland Kohring, Leila Lo Leggio
Bradyrhizobium japonicum sorbitol dehydrogenase is NADH-dependent and is active at elevated temperatures. The best substrate is D-glucitol (a synonym for D-sorbitol), although L-glucitol is also accepted, giving it particular potential in industrial applications. Crystallization led to a hexagonal crystal form, with crystals diffracting to 2.9 Å resolution. In attempts to phase the data, a molecular-replacement solution based upon PDB entry 4nbu (33% identical in sequence to the target) was found. The solution contained one molecule in the asymmetric unit, but a tetramer similar to that found in other short-chain dehydrogenases, including the search model, could be reconstructed by applying crystallographic symmetry operations...
November 1, 2016: Acta Crystallographica. Section F, Structural Biology Communications
R Zagrodnik, M Łaniecki
Hydrogen production from starch by a co-culture hybrid dark and photofermentation under repeated fed-batch conditions at different organic loading rates (OLR) was studied. Effective cooperation between bacteria in co-culture during initial days was observed at controlled pH 7.0. However, at pH above 6.5 dark fermentation phase was redirected from H2 formation towards production of formic acid, lactic acid and ethanol (which are not coupled with hydrogen production) with simultaneous lower starch removal efficiency...
October 21, 2016: Bioresource Technology
Lennart Weber, Clemens Thoelken, Marcel Volk, Bernhard Remes, Marcus Lechner, Gabriele Klug
Cell division and cell wall synthesis mechanisms are similarly conserved among bacteria. Consequently some bacterial species have comparable sets of genes organized in the dcw (division and cell wall) gene cluster. Dcw genes, their regulation and their relative order within the cluster are outstandingly conserved among rod shaped and gram negative bacteria to ensure an efficient coordination of growth and division. A well studied representative is the dcw gene cluster of E. coli. The first promoter of the gene cluster (mraZ1p) gives rise to polycistronic transcripts containing a 38 nt long 5' UTR followed by the first gene mraZ...
2016: PloS One
Florian Richter, Ines Fonfara, Boris Bouazza, Charlotte Helene Schumacher, Majda Bratovič, Emmanuelle Charpentier, Andreas Möglich
Sensory photoreceptors have enabled non-invasive and spatiotemporal control of numerous biological processes. Photoreceptor engineering has expanded the repertoire beyond natural receptors, but to date no generally applicable strategy exists towards constructing light-regulated protein actuators of arbitrary function. We hence explored whether the homodimeric Rhodobacter sphaeroides light-oxygen-voltage (LOV) domain (RsLOV) that dissociates upon blue-light exposure can confer light sensitivity onto effector proteins, via a mechanism of light-induced functional site release...
November 16, 2016: Nucleic Acids Research
Benjamin W Arentson, Erin L Hayes, Weidong Zhu, Harkewal Singh, John J Tanner, Donald F Becker
Proline utilization A (PutA) is a bifunctional flavoenzyme with proline dehydrogenase (PRODH) and Δ(1)-pyrroline-5-carboxylate dehydrogenase domains that catalyzes the two-step oxidation of proline to glutamate. Trifunctional PutAs also have an N-terminal ribbon-helix-helix (RHH) DNA-binding domain and moonlight as autogenous transcriptional repressors of the put regulon. A unique property of trifunctional PutA is the ability to switch functions from DNA-bound repressor to membrane-associated enzyme in response to cellular nutritional needs and proline availability...
October 14, 2016: Bioscience Reports
Andrea von Zadow, Elisabeth Ignatz, Richard Pokorny, Lars-Oliver Essen, Gabriele Klug
Photolyases are efficient DNA repair enzymes that specifically repair either cyclobutane pyrimidine dimers or (6-4) photoproducts in a light-dependent cleavage reaction. The closely related classical cryptochrome blue light photoreceptors do not repair DNA lesions; instead they are involved in regulatory processes. CryB of Rhodobacter sphaeroides was until now described as a cryptochrome that affects light-dependent and singlet oxygen-dependent gene expression and is unusual in terms of its cofactor composition...
December 2016: FEBS Journal
Cosima Damiana Calvano, Giovanni Ventura, Massimo Trotta, Giuliana Bianco, Tommaso R I Cataldi, Francesco Palmisano
Bacteriochlorophyll a (BChl a), a photosynthetic pigment performing the same functions of chlorophylls in plants, features a bacteriochlorin macrocycle ring (18 π electrons) with two reduced pyrrole rings along with a hydrophobic terpenoid side chain (i.e., the phytol residue). Chlorophylls analysis by matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) is not so straightforward since pheophytinization (i.e., release of the central metal ion) and cleavage of the phytol-ester linkage are invariably observed by employing protonating matrices such as 2,5-dihydroxybenzoic acid, sinapinic acid, and α-cyano-4-hydroxycinnamic acid...
October 11, 2016: Journal of the American Society for Mass Spectrometry
Nikolaos Remmas, Paraschos Melidis, Chrysoula Voltsi, Despoina Athanasiou, Spyridon Ntougias
Mature landfill leachate is a heavily-polluted wastewater due to its recalcitrant nature of organic matter, and high ammonia and salt content. Despite the moderate saline and alkaline nature of this habitat, no attention has been paid to the isolation and functional role of extremophiles in such environment. In this work, a total of 73 and 29 bacterial strains were isolated by using alkaline and saline media, respectively, while bacteria from mature landfill leachate growing in these media were enumerated as 1...
October 11, 2016: Journal of Environmental Science and Health. Part A, Toxic/hazardous Substances & Environmental Engineering
Lilit Gabrielyan, Lilit Hakobyan, Armen Trchounian
The aim of the present work was the study of comparative effects of Cu(II) and Ni(II) and their mixture on growth, redox potential, hydrogen (H2) yield and ATPase activity in phototrophic purple bacteria R. sphaeroides MDC6522 from Jermuk mineral spring in Armenia. It was ascertained, that Cu(2+) and Ni(2+) have different effects on bacterial specific growth rate: in the presence of 5μM Cu(2+) growth rate was ~3.2-fold lower in comparison with control (no addition), and increased ~1.5-fold in medium with 5μM Ni(2+)...
October 1, 2016: Journal of Photochemistry and Photobiology. B, Biology
Shan Wen, Hongyu Liu, Huijun He, Le Luo, Xiang Li, Guangming Zeng, Zili Zhou, Wei Lou, Chunping Yang
Two strains of photosynthetic bacteria, Rhodobacter blasticus and Rhodobacter capsulatus, were used in this work to investigate the feasibility of using photosynthetic bacteria for the treatment of anaerobically digested swine wastewater. The effects of crucial factors which influence the pollutants removal efficiency were also examined. Results showed that anaerobically digested swine wastewater could be treated effectively by photosynthetic bacteria. The treatment efficiency was significantly higher by the mixed photosynthetic bacteria than that by any unitary bacterium...
September 30, 2016: Bioresource Technology
Lijun Xi, Nenghu Qiao, Zhenchong Zhang, Laihong Yan, Fengbo Li, Jinpeng Hu, Jing Li
A novel bacterium, strain L3(T), was isolated from an activated sludge sample retrieved from a municipal wastewater treatment plant in Huangdao, China. On the basis of 16S rRNA gene sequence similarity studies, strain L3(T) was affiliated to the genus Sinorhodobacter, being most closely related to Sinorhodobacter ferrireducens (98.0 %). The 16S rRNA gene sequence similarity of strain L3(T) to other related species, Thioclava atlantica DLFJ1-1(T) (96.5 %), Rhodobacter capsulatus ATCC 11166(T) (96.3 %), Paenirhodobacter enshiensis DW2-9(T) (96...
September 30, 2016: Antonie Van Leeuwenhoek
Anna Tsargorodska, Michaël L Cartron, Cvetelin Vasilev, Goutham Kodali, Olga A Mass, Jeremy J Baumberg, P Leslie Dutton, C Neil Hunter, Päivi Törmä, Graham J Leggett
Gold nanostructure arrays exhibit surface plasmon resonances that split after attaching light harvesting complexes 1 and 2 (LH1 and LH2) from purple bacteria. The splitting is attributed to strong coupling between the localized surface plasmon resonances and excitons in the light-harvesting complexes. Wild-type and mutant LH1 and LH2 from Rhodobacter sphaeroides containing different carotenoids yield different splitting energies, demonstrating that the coupling mechanism is sensitive to the electronic states in the light harvesting complexes...
October 10, 2016: Nano Letters
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