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J. Amesz - One of the best experts on this subject based on the ideXlab platform.
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electron transfer in reaction center core complexes from the Green Sulfur Bacteria prosthecochloris aestuarii and chlorobium tepidum
Biochemistry, 2000Co-Authors: Kristiane A Schmidt, Christine Hagerbraun, Sieglinde Neerken, Hjalmar P Permentier, J. AmeszAbstract:Electron transfer in reaction center core (RCC) complexes from the Green Sulfur Bacteria Prosthecochloris aestuarii and Chlorobium tepidum was studied by measuring flash-induced absorbance changes. The first preparation contained approximately three iron-Sulfur centers, indicating that the three putative electron acceptors F(X), F(A), and F(B) were present; the Chl. tepidum complex contained on the average only one. In the RCC complex of Ptc. aestuarii at 277 K essentially all of the oxidized primary donor (P840(+)) created by a flash was rereduced in several seconds by N-methylphenazonium methosulfate. In RCC complexes of Chl. tepidum two decay components, one of 0.7 ms and a smaller one of about 2 s, with identical absorbance difference spectra were observed. The fast component might be due to a back reaction of P840(+) with a reduced electron acceptor, in agreement with the notion that the terminal electron acceptors, F(A) and F(B), were lost in most of the Chl. tepidum complexes. In both complexes the terminal electron acceptor (F(A) or F(B)) could be reduced by dithionite, yielding a back reaction of 170 ms with P840(+). At 10 K in the RCC complexes of both species P840(+) was rereduced in 40 ms, presumably by a back reaction with F(X)(-). In addition, a 350 micros component occurred that can be ascribed to decay of the triplet of P840, formed in part of the complexes. For P840(+) rereduction a pronounced temperature dependence was observed, indicating that electron transfer is blocked after F(X) at temperatures below 200 K.
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Composition and optical properties of reaction centre core complexes from the Green Sulfur Bacteria Prosthecochloris aestuarii and Chlorobium tepidum
Photosynthesis Research, 2000Co-Authors: Hjalmar P Permentier, Masami Kobayashi, Kristiane A Schmidt, Sieglinde Neerken, Mette Miller, Machiko Akiyama, Christine Hager-braun, J. AmeszAbstract:Photosynthetically active reaction centre core (RCC) complexes were isolated from two species of Green Sulfur Bacteria, Prosthecochloris (Ptc.) aestuarii strain 2K and Chlorobium ( Chl .) tepidum , using the same isolation procedure. Both complexes contained the main reaction centre protein PscA and the iron–Sulfur protein PscB, but were devoid of Fenna–Matthews–Olson (FMO) protein. The Chl. tepidum RCC preparation contained in addition PscC (cytochrome c ). In order to allow accurate determination of the pigment content of the RCC complexes, the extinction coefficients of bacteriochlorophyll (BChl) a in several solvents were redetermined with high precision. They varied between 54.8 mM^−1 cm^−1 for methanol and 97.0 mM^−1 cm^−1 for diethylether in the Q_Y maximum. Both preparations appeared to contain 16 BChls a of which two are probably the 13^2-epimers, 4 chlorophylls (Chls) a 670 and 2 carotenoids per RCC. The latter were of at least two different types. Quinones were virtually absent. The absorption spectra were similar for the two species, but not identical. Eight bands were present at 6 K in the BChl a Q_Y region, with positions varying from 777 to 837 nm. The linear dichroism spectra showed that the orientation of the BChl a Q_Y transitions is roughly parallel to the membrane plane; most nearly parallel were transitions at 800 and 806 nm. For both species, the circular dichroism spectra were dominated by a strong band at 807–809 nm, indicating strong interactions between at least some of the BChls. The absorption, CD and LD spectra of the four Chls a 670 were virtually identical for both RCC complexes, indicating that their binding sites are highly conserved and that they are an essential part of the RCC complexes, possibly as components of the electron transfer chain. Low temperature absorption spectroscopy indicated that typical FMO–RCC complexes of Ptc. aestuarii and Chl. tepidum contain two FMO trimers per reaction centre.
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the primary electron acceptor of Green Sulfur Bacteria bacteriochlorophyll 663 is chlorophyll a esterified with δ 2 6 phytadienol
Photosynthesis Research, 2000Co-Authors: Masami Kobayashi, J. Amesz, Tadashi Watanabe, Hirozo Ohoka, Satoshi Akutsu, Machiko Akiyama, Keisuke Tominaga, Hideo Kise, Fumiko Nishida, Mika KoizumiAbstract:The primary electron acceptor of Green Sulfur Bacteria, bacteriochlorophyll (BChl) 663, was isolated at high purity by an improved purification procedure from a crude reaction center complex, and the molecular structure was determined by fast atom bombardment mass spectroscopy (FAB-mass), 1H- and 13C-NMR spectrometry, double quantum filtered correlation spectroscopy (DQF-COSY), heteronuclear multiple-quantum coherence (HMQC) and heteronuclear multiple-bond correlation (HMBC) spectral measurements. BChl 663 was 2.0 mass units smaller than plant Chl a. The NMR spectra showed that the macrocycle was identical to that of Chl a. In the esterifying alcohol, a singlet P71 signal was observed at the high-field side of the singlet P31 signal in BChl 663, while a doublet peak of P71 overlapped that of P111 in Chl a. A signal of P7-proton, seen in Chl a, was lacking, and the P6-proton appeared as a triplet signal near the triplet P2-proton signal in BChl 663. These results indicate the presence in BChl 663 of a C=C double bond between P6 and P7 in addition to that between P2 and P3. The structure of BChl 663 was hence concluded to be Chl a esterified with 2,6-phytadienol instead of phytol. In addition to BChl 663, two molecules of the 132-epimer of BChl a, BChl a′, were found to be present per reaction center, which may constitute the primary electron donor.
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excited state dynamics in fmo antenna complexes from photosynthetic Green Sulfur Bacteria a kinetic model
Journal of Physical Chemistry B, 1999Co-Authors: S I E Vulto, J. Amesz, Sieglinde Neerken, Michiel A De Baat, Frank R Nowak, H Van Amerongen, Thijs J AartsmaAbstract:We have simulated the excited state dynamics of the FMO (Fenna−Matthews−Olson) bacteriochlorophyll a-protein complexes of the Green Sulfur Bacteria Chlorobium (C.) tepidum and Prosthecochloris (P.) aestuarii at cryogenic temperature in terms of an exciton model. The simulation is based on the electronic structure, as described in previous publications (Louwe, R. J. W.; Vrieze, J.; Hoff, A. J.; Aartsma, T. J.; J. Phys. Chem. 1997, 101, 11280. Vulto, S. I. E.; de Baat, M. A.; Louwe, R. J. W.; Permentier, H. P.; Neef, T.; Miller, M.; van Amerongen, H.; Aartsma, T. J. J. Phys. Chem. 1998, 102, 9577). Relaxation between exciton states is described by linear electron phonon coupling as a perturbation term in the Hamiltonian of the system. The simulation was compared with experimental data obtained by pump−probe measurements with various wavelengths of excitation. For C. tepidum, a quite good agreement was obtained between the calculated and measured dynamics. For P. aestuarii, the simulations are less satisfact...
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isolation and pigment composition of the antenna system of four species of Green Sulfur Bacteria
Photosynthesis Research, 1997Co-Authors: Christof Francke, J. AmeszAbstract:New and rapid procedures were developed for the isolation of chlorosomes and FMO-protein from the Green Sulfur Bacteria Prosthecochloris (P.) aestuarii, Chlorobium (Cb.) phaeovibrioides, Cb. tepidum and Cb. vibrioforme. The resulting preparations were free from contaminating pigments and proteins as was shown by absorption spectroscopy, pigment analysis and SDS-PAGE. Two spectrally different types of FMO-protein were found. The first type, present in P. aestuarii and Cb. vibrioforme, has a main absorption band at 6 K at 815 nm, whereas the second type, isolated from Cb. tepidum and Cb. phaeovibrioides, has a strong band at 806 nm. In contrast to what was recently suggested (Tronrud DE and Matthews BW (1993) In: Deisenhofer J and Norris J (eds) The Photosynthetic Reaction Center, Vol 1, pp 13–21. Academic Press, San Diego, CA) the FMO-proteins contained no polar BChl a homologue. The isolated chlorosomes showed a small blue-shift of the QY absorption maximum with respect to intact cells. For the different species, grown under the same light conditions, the homologue composition of BChls c and d was approximately identical whereas for the BChl e in Cb. phaeovibrioides the relative amounts of homologues with larger alkyl substituents at position 8 were considerably larger. Baseplate BChl a was present in all chlorosomes and comprised 1–2% of the chlorosomal BChl. Its QY absorption band was located at about 802 nm and was clearly separated from the major QY absorption band at 6 K. The predominant esterifying alcohol of BChl a in the chlorosomes as well as in the FMO-proteins was phytol, but both antenna complexes also contained small amounts of BChl a esterified with the metabolic intermediates geranylgeraniol, dihydrogeranylgeraniol and tetrahydrogeranylgeraniol, like most purple Bacteria. Since the esterifying alcohols of the chlorosomal BChl a and of the main chlorosomal pigments (BChls c, d and e) are different, esterification, and perhaps also the synthesis, of the BChls in the interior of the chlorosome and of the BChls in the baseplate must be spatially and genetically separated processes.
Donald A. Bryant - One of the best experts on this subject based on the ideXlab platform.
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Host population diversity as a driver of viral infection cycle in wild populations of Green Sulfur Bacteria with long standing virus-host interactions
The ISME Journal, 2021Co-Authors: Maureen Berg, Jennifer L. Thweatt, Donald A. Bryant, Danielle Goudeau, Charles Olmsted, Katherine D. Mcmahon, Senay Yitbarek, Emiley A. Eloe-fadrosh, Rex R. Malmstrom, Simon RouxAbstract:Temperate phages are viruses of Bacteria that can establish two types of infection: a lysogenic infection in which the virus replicates with the host cell without producing virions, and a lytic infection where the host cell is eventually destroyed, and new virions are released. While both lytic and lysogenic infections are routinely observed in the environment, the ecological and evolutionary processes regulating these viral dynamics are still not well understood, especially for uncultivated virus-host pairs. Here, we characterized the long-term dynamics of uncultivated viruses infecting Green Sulfur Bacteria (GSB) in a model freshwater lake (Trout Bog Lake, TBL). As no GSB virus has been formally described yet, we first used two complementary approaches to identify new GSB viruses from TBL; one in vitro based on flow cytometry cell sorting, the other in silico based on CRISPR spacer sequences. We then took advantage of existing TBL metagenomes covering the 2005–2018 period to examine the interactions between GSB and their viruses across years and seasons. From our data, GSB populations in TBL were constantly associated with at least 2-8 viruses each, including both lytic and temperate phages. The dominant GSB population in particular was consistently associated with two prophages with a nearly 100% infection rate for >10 years. We illustrate with a theoretical model that such an interaction can be stable given a low, but persistent, level of prophage induction in low-diversity host populations. Overall, our data suggest that lytic and lysogenic viruses can readily co-infect the same host population, and that host strain-level diversity might be an important factor controlling virus-host dynamics including lytic/lysogeny switch.
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host population diversity as a driver of viral infection cycle in wild populations of Green Sulfur Bacteria with long standing virus host interactions
bioRxiv, 2020Co-Authors: Maureen Berg, Jennifer L. Thweatt, Donald A. Bryant, Danielle Goudeau, Charles Olmsted, Katherine D. Mcmahon, Rex R. Malmstrom, Emiley A Eloefadrosh, Simon RouxAbstract:ABSTRACT Viral infections of Bacterial hosts range from highly lytic to lysogenic, where highly lytic viruses undergo viral replication and immediately lyse their hosts, and lysogenic viruses have a latency period before replication and host lysis. While both types of infections are routinely observed in the environment, the ecological and evolutionary processes that regulate these different viral dynamics are still not well understood. In this study, we identify and characterize the long-term dynamics of uncultivated viruses infecting Green Sulfur Bacteria (GSB) in a model freshwater lake sampled from 2005-2018. Because of the additional requirements for the laboratory cultivation of anaerobes like GSB, viruses infecting GSB have yet to be formally identified, leaving their diversity and impact on natural populations of GSB virtually unknown. In this study, we used two approaches to identify viruses infecting GSB; one in vitro based on flow cytometry cell sorting, the other in silico based on CRISPR spacer sequences. We then took advantage of existing bulk metagenomes derived from Trout Bog Lake covering the 2005-2018 period to examine the interactions between GSB hosts and their viruses across multiple years and seasons. From our data, GSB populations in Trout Bog Lake were found to be concurrently infected with at least 2-8 viruses each, many of which were lysogenic viruses; one GSB host population in particular was consistently associated with two lysogens with a nearly 100% infection rate for over 10 years. We illustrate with a theoretical infection model that such an interaction can be stable over multiple years given a low, but persistent level of lysogen induction in host populations with already high infection rates. Overall, our data suggest that single GSB populations are typically infected by multiple viruses at the same time, that lytic and lysogenic viruses can readily co-infect the same host population in the same ecosystem, and that host strain-level diversity might be an important factor controlling the lytic/lysogeny switch.
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A paralog of a bacteriochlorophyll biosynthesis enzyme catalyzes the formation of 1,2-dihydrocarotenoids in Green Sulfur Bacteria
The Journal of biological chemistry, 2018Co-Authors: Daniel P. Canniffe, C. Neil Hunter, Jennifer L. Thweatt, Aline Gomez Maqueo Chew, Donald A. BryantAbstract:Chlorobaculum tepidum, a Green Sulfur bacterium, utilizes chlorobactene as its major carotenoid, and this organism also accumulates a reduced form of this monocyclic pigment, 1',2'-dihydrochlorobactene. The protein catalyzing this reduction is the last unidentified enzyme in the biosynthetic pathways for all of the Green Sulfur Bacterial pigments used for photosynthesis. The genome of Chlorobaculum tepidum contains two paralogous genes encoding members of the FixC family of flavoproteins: bchP, that has been shown to encode an enzyme of bacteriochlorophyll biosynthesis; and bchO, for which a function has not been assigned. Here we demonstrate that a bchO mutant is unable to synthesize 1',2'-dihydrochlorobactene, and when bchO is heterologously expressed in a neurosporene-producing mutant of the purple bacterium, Rhodobactersphaeroides, the encoded protein is able to catalyze the formation of 1,2-dihydroneurosporene, the major carotenoid of the only other organism reported to synthesize 1,2-dihydrocarotenoids, Blastochloris viridis Identification of this enzyme completes the pathways for the synthesis of photosynthetic pigments in Chlorobiaceae, and accordingly and consistent with its role in carotenoid biosynthesis, we propose to rename the gene, cruI Notably, the absence of cruI in Blastochloris viridis indicates that a second 1,2-carotenoid reductase, which is structurally unrelated to CruI (BchO), must exist in nature. The evolution of this carotenoid reductase in Green Sulfur Bacteria is discussed herein.
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speciation and ecological success in dimly lit waters horizontal gene transfer in a Green Sulfur Bacteria bloom unveiled by metagenomic assembly
The ISME Journal, 2017Co-Authors: Tomas Llorensmares, Donald A. Bryant, Zhenfeng Liu, Lisa Zeigler Allen, Douglas B Rusch, Matthew T Craig, Christopher L Dupont, Emilio O CasamayorAbstract:A natural planktonic bloom of a brown-pigmented photosynthetic Green Sulfur Bacteria (GSB) from the disphotic zone of karstic Lake Banyoles (NE Spain) was studied as a natural enrichment culture from which a nearly complete genome was obtained after metagenomic assembly. We showed in situ a case where horizontal gene transfer (HGT) explained the ecological success of a natural population unveiling ecosystem-specific adaptations. The uncultured brown-pigmented GSB was 99.7% identical in the 16S rRNA gene sequence to its Green-pigmented cultured counterpart Chlorobium luteolum DSM 273T. Several differences were detected for ferrous iron acquisition potential, ATP synthesis and gas vesicle formation, although the most striking trait was related to pigment biosynthesis strategy. Chl. luteolum DSM 273T synthesizes bacteriochlorophyll (BChl) c, whereas Chl. luteolum CIII incorporated by HGT a 18-kbp cluster with the genes needed for BChl e and specific carotenoids biosynthesis that provided ecophysiological advantages to successfully colonize the dimly lit waters. We also genomically characterized what we believe to be the first described GSB phage, which based on the metagenomic coverage was likely in an active state of lytic infection. Overall, we observed spread HGT and we unveiled clear evidence for virus-mediated HGT in a natural population of photosynthetic GSB.
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multiple types of 8 vinyl reductases for bacterio chlorophyll biosynthesis occur in many Green Sulfur Bacteria
Journal of Bacteriology, 2011Co-Authors: Zhenfeng Liu, Donald A. BryantAbstract:Two 8-vinyl reductases, BciA and BciB, have been identified in chlorophototrophs. The bciA gene of Chlorobaculum tepidum was replaced with genes similar to bciB from other Green Sulfur Bacteria. Pigment analyses of the complemented strains showed that the bciB homologs encode 8-vinyl reductases similar to those of cyanoBacteria.
Kenichiro Suzuki - One of the best experts on this subject based on the ideXlab platform.
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ignavibacterium album gen nov sp nov a moderately thermophilic anaerobic bacterium isolated from microbial mats at a terrestrial hot spring and proposal of ignaviBacteria classis nov for a novel lineage at the periphery of Green Sulfur Bacteria
International Journal of Systematic and Evolutionary Microbiology, 2010Co-Authors: Koji Mori, Yoshihito Uchino, Tatsunori Nakagawa, Shigeaki Harayama, Kenichiro SuzukiAbstract:A moderately thermophilic chemoheterotrophic bacterium, strain Mat9-16(T), was isolated from microbial mats developed in hot spring water streams from Yumata, Nagano, Japan. Cells of strain Mat9-16(T) were strictly anaerobic, Gram-stain-negative, non-sporulating, non-motile and short to long rods (2.0-15.5 mum in length). Strain Mat9-16(T) grew fermentatively with optimum growth at 45 degrees C, pH 7.0-7.5 and 1 % NaCl (w/v). Phylogenetic analysis based on the 16S rRNA gene revealed that strain Mat9-16(T) was affiliated with an uncultivated lineage, and the nearest cultivated neighbours were Green Sulfur Bacteria belonging to the class Chlorobea with 77-83 % sequence similarity. However, strain Mat9-16(T) could not grow phototrophically and did not possess light-harvesting structures, morphologically and genetically, such as the chlorosomes of Green Sulfur Bacteria. On the basis of phenotypic features and phylogenetic position, a novel genus and species are proposed for strain Mat9-16(T), to be named Ignavibacterium album gen. nov., sp. nov. (=NBRC 101810(T) =DSM 19864(T)). We also propose to place the cultivated Bacterial lineage accommodating the sole representative Mat9-16(T) in a novel class, IgnaviBacteria classis nov. In addition, we present a formal description of the phylum-level taxon 'Chlorobi' as Chlorobi phyl. nov.
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ignavibacterium album gen nov sp nov a moderately thermophilic anaerobic bacterium isolated from microbial mats at a terrestrial hot spring and proposal of ignaviBacteria classis nov for a novel lineage at the periphery of Green Sulfur Bacteria
International Journal of Systematic and Evolutionary Microbiology, 2010Co-Authors: Koji Mori, Yoshihito Uchino, Tatsunori Nakagawa, Shigeaki Harayama, Kenichiro SuzukiAbstract:A moderately thermophilic chemoheterotrophic bacterium, strain Mat9-16T, was isolated from microbial mats developed in hot spring water streams from Yumata, Nagano, Japan. Cells of strain Mat9-16T were strictly anaerobic, Gram-stain-negative, non-sporulating, non-motile and short to long rods (2.0–15.5 μm in length). Strain Mat9-16T grew fermentatively with optimum growth at 45 °C, pH 7.0–7.5 and 1 % NaCl (w/v). Phylogenetic analysis based on the 16S rRNA gene revealed that strain Mat9-16T was affiliated with an uncultivated lineage, and the nearest cultivated neighbours were Green Sulfur Bacteria belonging to the class Chlorobea with 77–83 % sequence similarity. However, strain Mat9-16T could not grow phototrophically and did not possess light-harvesting structures, morphologically and genetically, such as the chlorosomes of Green Sulfur Bacteria. On the basis of phenotypic features and phylogenetic position, a novel genus and species are proposed for strain Mat9-16T, to be named Ignavibacterium album gen. nov., sp. nov. (=NBRC 101810T =DSM 19864T). We also propose to place the cultivated Bacterial lineage accommodating the sole representative Mat9-16T in a novel class, IgnaviBacteria classis nov. In addition, we present a formal description of the phylum-level taxon ‘Chlorobi’ as Chlorobi phyl. nov.
Koji Mori - One of the best experts on this subject based on the ideXlab platform.
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ignavibacterium album gen nov sp nov a moderately thermophilic anaerobic bacterium isolated from microbial mats at a terrestrial hot spring and proposal of ignaviBacteria classis nov for a novel lineage at the periphery of Green Sulfur Bacteria
International Journal of Systematic and Evolutionary Microbiology, 2010Co-Authors: Koji Mori, Yoshihito Uchino, Tatsunori Nakagawa, Shigeaki Harayama, Kenichiro SuzukiAbstract:A moderately thermophilic chemoheterotrophic bacterium, strain Mat9-16(T), was isolated from microbial mats developed in hot spring water streams from Yumata, Nagano, Japan. Cells of strain Mat9-16(T) were strictly anaerobic, Gram-stain-negative, non-sporulating, non-motile and short to long rods (2.0-15.5 mum in length). Strain Mat9-16(T) grew fermentatively with optimum growth at 45 degrees C, pH 7.0-7.5 and 1 % NaCl (w/v). Phylogenetic analysis based on the 16S rRNA gene revealed that strain Mat9-16(T) was affiliated with an uncultivated lineage, and the nearest cultivated neighbours were Green Sulfur Bacteria belonging to the class Chlorobea with 77-83 % sequence similarity. However, strain Mat9-16(T) could not grow phototrophically and did not possess light-harvesting structures, morphologically and genetically, such as the chlorosomes of Green Sulfur Bacteria. On the basis of phenotypic features and phylogenetic position, a novel genus and species are proposed for strain Mat9-16(T), to be named Ignavibacterium album gen. nov., sp. nov. (=NBRC 101810(T) =DSM 19864(T)). We also propose to place the cultivated Bacterial lineage accommodating the sole representative Mat9-16(T) in a novel class, IgnaviBacteria classis nov. In addition, we present a formal description of the phylum-level taxon 'Chlorobi' as Chlorobi phyl. nov.
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ignavibacterium album gen nov sp nov a moderately thermophilic anaerobic bacterium isolated from microbial mats at a terrestrial hot spring and proposal of ignaviBacteria classis nov for a novel lineage at the periphery of Green Sulfur Bacteria
International Journal of Systematic and Evolutionary Microbiology, 2010Co-Authors: Koji Mori, Yoshihito Uchino, Tatsunori Nakagawa, Shigeaki Harayama, Kenichiro SuzukiAbstract:A moderately thermophilic chemoheterotrophic bacterium, strain Mat9-16T, was isolated from microbial mats developed in hot spring water streams from Yumata, Nagano, Japan. Cells of strain Mat9-16T were strictly anaerobic, Gram-stain-negative, non-sporulating, non-motile and short to long rods (2.0–15.5 μm in length). Strain Mat9-16T grew fermentatively with optimum growth at 45 °C, pH 7.0–7.5 and 1 % NaCl (w/v). Phylogenetic analysis based on the 16S rRNA gene revealed that strain Mat9-16T was affiliated with an uncultivated lineage, and the nearest cultivated neighbours were Green Sulfur Bacteria belonging to the class Chlorobea with 77–83 % sequence similarity. However, strain Mat9-16T could not grow phototrophically and did not possess light-harvesting structures, morphologically and genetically, such as the chlorosomes of Green Sulfur Bacteria. On the basis of phenotypic features and phylogenetic position, a novel genus and species are proposed for strain Mat9-16T, to be named Ignavibacterium album gen. nov., sp. nov. (=NBRC 101810T =DSM 19864T). We also propose to place the cultivated Bacterial lineage accommodating the sole representative Mat9-16T in a novel class, IgnaviBacteria classis nov. In addition, we present a formal description of the phylum-level taxon ‘Chlorobi’ as Chlorobi phyl. nov.
Hirozo Oh-oka - One of the best experts on this subject based on the ideXlab platform.
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Gene expression system in Green Sulfur Bacteria by conjugative plasmid transfer.
PloS one, 2013Co-Authors: Chihiro Azai, Jiro Harada, Hirozo Oh-okaAbstract:Gene transfer and expression systems in Green Sulfur Bacteria were established by Bacterial conjugation with Escherichia coli. Conjugative plasmid transfer from E. coli S17-1 to a thermophilic Green Sulfur bacterium, Chlorobaculum tepidum (formerly Chlorobium tepidum) WT2321, was executed with RSF1010-derivative broad-host-range plasmids, named pDSK5191 and pDSK5192, that confer erythromycin and streptomycin/spectinomycin resistance, respectively. The transconjugants harboring these plasmids were reproducibly obtained at a frequency of approximately 10-5 by selection with erythromycin and a combination of streptomycin and spectinomycin, respectively. These plasmids were stably maintained in C. tepidum cells in the presence of these antibiotics. The plasmid transfer to another mesophilic Green Sulfur bacterium, C. limnaeum (formerly Chlorobium phaeobacteroides) RK-j-1, was also achieved with pDSK5192. The expression plasmid based on pDSK5191 was constructed by incorporating the upstream and downstream regions of the pscAB gene cluster on the C. tepidum genome, since these regions were considered to include a constitutive promoter and a ρ-independent terminator, respectively. Growth defections of the ∆cycA and ∆soxB mutants were completely rescued after introduction of pDSK5191-cycA and -soxB that were designed to express their complementary genes. On the other hand, pDSK5191-6xhis-pscAB, which incorporated the gene cluster of 6xhis-pscA and pscB, produced approximately four times more of the photosynthetic reaction center complex with His-tagged PscA as compared with that expressed in the genome by the conventional natural transformation method. This expression system, based on conjugative plasmid, would be applicable to general molecular biological studies of Green Sulfur Bacteria.
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Gene Expression System in Green Sulfur Bacteria by Conjugative Plasmid Transfer
2013Co-Authors: Chihiro Azai, Jiro Harada, Hirozo Oh-okaAbstract:Gene transfer and expression systems in Green Sulfur Bacteria were established by Bacterial conjugation with Escherichia coli. Conjugative plasmid transfer from E. coli S17-1 to a thermophilic Green Sulfur bacterium, Chlorobaculum tepidum (formerly Chlorobium tepidum) WT2321, was executed with RSF1010-derivative broad-host-range plasmids, named pDSK5191 and pDSK5192, that confer erythromycin and streptomycin/spectinomycin resistance, respectively. The transconjugants harboring these plasmids were reproducibly obtained at a frequency of approximately 10-5 by selection with erythromycin and a combination of streptomycin and spectinomycin, respectively. These plasmids were stably maintained in C. tepidum cells in the presence of these antibiotics. The plasmid transfer to another mesophilic Green Sulfur bacterium, C. limnaeum (formerly Chlorobium phaeobacteroides) RK-j-1, was also achieved with pDSK5192. The expression plasmid based on pDSK5191 was constructed by incorporating the upstream and downstream regions of the pscAB gene cluster on the C. tepidum genome, since these regions were considered to include a constitutive promoter and a ρ-independent terminator, respectively. Growth defections of the ∆cycA and ∆soxB mutants were completely rescued after introduction of pDSK5191-cycA and-soxB that were designed to express their complementary genes. On the other hand, pDSK5191-6xhis-pscAB, which incorporated the gene cluster of 6xhis-pscA and pscB, produced approximately four times more of th
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C-type cytochromes in the photosynthetic electron transfer pathways in Green Sulfur Bacteria and helioBacteria
Photosynthesis Research, 2010Co-Authors: Chihiro Azai, Shigeru Itoh, Hirozo Oh-okaAbstract:Green Sulfur Bacteria and helioBacteria are strictly anaerobic phototrophs that have homodimeric type 1 reaction center complexes. Within these complexes, highly reducing substances are produced through an initial charge separation followed by electron transfer reactions driven by light energy absorption. In order to attain efficient energy conversion, it is important for the photooxidized reaction center to be rapidly rereduced. Green Sulfur Bacteria utilize reduced inorganic Sulfur compounds (sulfide, thiosulfate, and/or Sulfur) as electron sources for their anoxygenic photosynthetic growth. Membrane-bound and soluble cytochromes c play essential roles in the supply of electrons from Sulfur oxidation pathways to the P840 reaction center. In the case of gram-positive helioBacteria, the photooxidized P800 reaction center is rereduced by cytochrome c -553 (PetJ) whose N-terminal cysteine residue is modified with fatty acid chains anchored to the cytoplasmic membrane.
