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Fuminori Takahashi, Tsuyoshi Mizoguchi, Riichiro Yoshida, Kazuya Ichim ...
Pages
0551
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
FREE ACCESS
Mitogen-activated protein kinase (MAPK) cascades are universal signal transduction modules in eukaryotes. In Arabidopsis genome, there are 20 genes encoding possible MAPKs, and these MAPKs can be divided into at least four groups (A-D). Group D has 8 members, and forms the largest subgroup. Group D MAPKs contain TDY motifs instead of TEY motifs in their T-loop and larger C-terminal regions relative to other groups. We have focused our work on MPK8, one of the group D MAPK, for further functional analysis. Yeast two-hybrid and
Arabidopsis protoplast BiFC analysis revealed that MPK8 interacts with various types of calmodulin. We have shown that calmodulin can activate MPK8 without calcium
in vitro. These results suggest a novel activation mechanism of MAPK by calmodulin
in plant. We will discuss the activation mechanism of MPK8.
View full abstract
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Akari Nakasone, Maki Kawai-Yamada, Issay Narumi, Hirofumi Uchimiya, Yu ...
Pages
0552
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
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The
Small Acidic Protein 1 (
SMAP1) gene was identified as the regulator for the 2,4-D response in
Arabidopsis root. SMAP1 has the conserved phenylalanine and aspartate rich domain (F/D). Its orthologous genes are highly conserved in plants and animals, suggesting SMAP1 has significant biological functions. To investigate the functions of SMAP1,
SMAP1-GFP or
SMAP1 ΔF/D-GFP were introduced to the
aar1 mutant that is deficient in the
SMAP1 gene.
SMAP1-GFP recovered 2,4-D sensitivity of the
aar1 mutant while
SMAP1 ΔF/D-GFP did not, implying that F/D region is required for the SMAP1 functions. Next, using anti-GFP-MicroBeads, the proteins, which form complexes with SMAP1-GFP but not with SMAP1 ΔF/D-GFP, were pulled down and analyzed by MS. The result indicated that SMAP1-GFP binds to COP9 signalosome (CSN). The results implied that SMAP1 regulates 2,4-D response by interacting to CSN through F/D region.
View full abstract
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Mayuko Kataoka, Shiori Aki, Naoshi Dohmae, Atsuhiro Oka, Takashi Aoyam ...
Pages
0553
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
FREE ACCESS
COP9 signalosome is a protein complex known to regulate ubiquitin-mediated protein degradation. Independent of this proteolysis, CSN subunit1 (CSN1) regulates signal transduction, possibly through transcriptional repression. In plants, the N-terminal half of CSN1 (CSN1N) is essential for survival. In order to reveal regulatory mechanism of CSN1N at the molecular level, we aim to systematically identify CSN1N-binding factors in plants. To date, general regulators on mRNA processing have been identified in mammals, while specific transcription factors have been identified in
Arabidopsis. Systematic analysis on, CSN1-interacting factors should reveal the conservation and differentiation among the different kingdoms.
Here, we analyzed expression conditions of plant CSN1N
in vitro, when fused to various tags. Flag-CSN1N gave best results for expression quantity and quality. We then subjected Flag-CSN1N to immuno-precipitation experiments against protein extracted from
Arabidopsis. We will further discuss the details and results of this approach.
View full abstract
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Tatsuru Masuda, Shigekazu Takahashi, Koichi Kobayashi
Pages
0554
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
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Heme is an essential molecule for various biological functions. Recent studies also suggest that heme functions as organelle-derived signal that regulates fundamental cell processes. The classic method of measuring heme by pyridine hemochrome is time, labor, and material intensive, and therefore limiting in its utility. Recently, we reported alternative method that is sensitive and specific to heme, which is based on the ability of horseradish peroxidase (HRP) apo-enzyme to reconstitute with heme to form an active holo-enzyme. Here, we developed high-throughput heme assay by performing reactions on multi-well plate with highly sensitive chemiluminescence detection reagents. Detection of chemiluminescence in CCD-based gel doc apparatus enables simultaneous measurement of multiple samples. Furthermore, high sensitivity of this assay allowed a direct measurement of heme in solvent extracts after dilution. This assay is sensitive, quick, provides a large dynamic range and is well suited for large-scale analysis of heme extracted from minute samples.
View full abstract
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Rina Aoki, Takeaki Goto, Kei Minamizaki, Yuichi Fujita
Pages
0555
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
FREE ACCESS
Heme oxygenase (HO) catalyzes the oxygen-dependent cleavage of the porphyrin ring of heme, producing biliverdin IXα in the phycobilin biosynthesis. There are two isoforms of heme oxygenase encoded by
ho1 (
sll1184) and
ho2 (
sll1875) with 51% identity in amino acid sequence in the cyanobacterium
Synechocystis sp. PCC 6803. Both isoforms HO1 and HO2 have been demonstrated to show heme oxygenase activity. However, the functional differentiation of the two HOs in the cyanobacterium remains unknown. We tried to isolate mutants in which one of two genes
ho1 and
ho2 was inactivated. In this presentation we will discuss possible functional differentiation based on the observed phenotype of the two mutants.
View full abstract
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Keigo Hori, Nobuyoshi Mochizuki, Tatsuru Masuda
Pages
0556
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
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Mg-chelatase composed by CHLI, CHLD, and CHLH subunits catalyzes the insertion of Mg2+ into protoporphyrin IX, the first step of chlorophyll biosynthesis. CHLH is catalytic subunit that binds substrate or product porphyrin. Recently, CHLH is proposed to be a plastid-localized abscisic acid receptor, and furthermore, mutations of CHLH resulted in
gun phenotype that defects in plastid to nuclear signaling. Here, we analyzed porphyrin-binding properties of Arabidopsis CHLH expressed in
E. coli. Soluble homogenous CHLH was successfully obtained by low-temperature induction and affinity chromatography. Although no porphyrin was bound to the resultant CHLH, this protein bound to protoporphyrin IX and deuteroporphyrin IX. Red shifts in fluorescence excitation peaks were observed in the bound porphyrin, suggesting slightly deformed non-planar conformation of the porphyrin. The emission from Trp residues in CHLH was partly quenched when porphyrin was bound. Porphyrin-binding properties of CHLH mutants that resulted in gun phenotype will also be presented.
View full abstract
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Yasushi Kawajiri, Kei Minamizaki, Yuichi Fujita
Pages
0557
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
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Isocyclic ring, a unique feature of all chlorophylls, is formed by Mg-protoporphyrin IX monomethylester (MPE) cyclase. In our previous work, we found two oxygen-dependent MPE cyclase isoforms, ChlA
I and ChlA
II, in the cyanobacterium
Synechocystis sp. PCC 6803. The constitutive isoform ChlA
I is essential for growth under the aerobic conditions, and the induced isoform ChlA
II is the dominant MPE cyclase operating together with ChlA
I under anaerobic (micro-oxic) conditions. In this study we isolated a series of mutants in which
chlAI and
chlAII were replaced each other to examine the compatibility of the isoforms. While a mutant in which
chlAI was replaced with
chlAII grew as well as wild type under both aerobic and anaerobic conditions, another mutant in which
chlAII was replaced with
chlAI showed growth retardation similar to
ΔchlAII under anaerobic conditions. This result suggests that ChlA
II has some special enzymatic properties for low oxygen conditions where ChlA
I operates inadequately.
View full abstract
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Jiro Harada, Shohei Miyago, Tadashi Mizoguchi, Hitoshi Tamiaki, Hirozo ...
Pages
0558
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
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The green sulfur bacterium
Chlorobium tepidum contains bacteriochlorophyll (BChl)
aP and chlorophyll (Chl)
aPD as minor chlorophyllous components as well as major BChl
c. BChl
aP acts as antenna pigments and P840, and Chl
aPD serves as a primary acceptor in the reaction center. The former pigment possesses a phytyl group on the C-17 position, while the latter is esterified with delta-2,6-phytadienol. Although it is known that both long hydrocarbon chains are synthesized from a geranylgeranyl group by geranylgeranyl reductase, CT2256, it is not still revealed that why CT2256 synthesizes different long hydrocarbon chains in BChl
aP and Chl
aPD. To clarify the reaction process of CT2256, we constructed mutants of
Chlorobium tepidum, that were deleted
CT2256 gene, and then introduced either cyanobacterial
chlP gene or purple bacterial
bchP gene. We will discuss the reduction process of CT2256 from results of pigments analysis of above mutants.
View full abstract
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Momoko Sato, Rei Narikawa, Masahiko Ikeuchi
Pages
0559
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
FREE ACCESS
Phycobilisomes are supramolecular complexes of phycobiliproteins associated with linker proteins and serve as major light harvesting antennae in cyanobacteria, red algae and glaucophyta. To date, further fractionation gave subcomplexes such as phycocyanin and allophycocyanin disks and core membrane linker with allophycocyanins. Previously we reported a new type of linker proteins (CpcK1, CpcK2) in the phycocyanin rods prepared from the phycobilisome of a glaucocystophyte
Cyanophora paradoxa. Here we attempted to fractionate the phycobilisome by native PAGE and found novel large subcomplex that included CpcK1/2 proteins in addition to core membrane linker and allophycocyanins. Since only a small amount of phycocyanins remained in the subcomplex, it is proposed that the phycobilisome supercomplex can be assembled from the core part with the aid of rod linker proteins followed by integration of the phycocyanin disks onto the linker.
View full abstract
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Yoshiaki Nakagawa, Yukari Tahara, Yutaka Shibata, Sigeru Itoh
Pages
0560
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
FREE ACCESS
When etiolated leaves of angiosperms are exposed to light, light-dependent chlorophyll (Chl) synthesis is triggered by the photoreduction of protochlorophyllide (Pchlide) into Chl. Subsequent development of the photosynthetic apparatus is completed within a few hours or a day. Little is known about the assembly process of synthesized Chl to antenna apoproteins so far.
Here, we studied the fluorescence dynamics of leaves during greening over the whole spectral range of Chl at 77 K. Greening of dark-grown Zea mays for 7 days was initiated by the white light illumination. After 0.5, 1, 2, 3, 4, 5 hours, leaves were harvested and immersed in liquid nitrogen to prevent further greening. The analysis of the fluorescence dynamics of thus prepared samples showed that the energy transfers to photosystem I red Chl emerged about 2 hours after the initiation of greening.
We discuss the assembly process of the antenna system in the greening leaves.
View full abstract
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Koichi Kobayashi, Hidehiro Fukaki, Tatsuru Masuda
Pages
0561
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
FREE ACCESS
During photomorphogenesis, chloroplasts are differentiated with concomitant activation of chlorophyll biosynthesis. Previously, we showed that in Arabidopsis roots, the chlorophyll biosynthesis, as well as the chloroplast differentiation is positively and negatively regulated by cytokinin and auxin, respectively, showing that these phytohormones are involved in the coordinated control of chlorophyll biosynthesis via transcriptional regulation. Here, we analyzed the effects of these phytohormones on chlorophyll biosynthesis during photomorphogenesis in Arabidopsis etiolated seedlings. Mutant analyses revealed that during photomorphogenesis, chlorophyll accumulation in cotyledons increased in auxin-signaling mutants
axr2,
shy2 and
slr, while decreased in a cytokinin receptor mutant
ahk2ahk3 when compared with wild-type. Since these phytohormones also regulate photomorphogenic changes such as hypocotyl elongation, it is likely that auxin and cytokinin oppositely affect the regulation of chlorophyll biosynthesis in aerial tissues, which is highly correlated to the tissue development and the cell specificity.
View full abstract
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Hiroyuki Hasegawa, Hisashi Ito, Ryouichi Tanaka, Ayumi Tanaka
Pages
0562
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
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Many enzymes for chlorophyll degradation are induced during senescence to convert dangerous pigments to safe molecules. The activities of these enzymes should be regulated for the efficient degradation, however, the regulation mechanism is till unknown. We studied the mutant which accumulates chlorophyll degradation products in Arabidopsis. Green leaves of the mutant accumulated 7-hydroxymethyl chlorophyll
a which is an intermediate molecule of chlorophyll
b to chlorophyll
a conversion. When the mutant was subjected to dark treatment to induce senescence, pheophorbide
a, a degradation product of chlorophyll
a, was accumulated. High-light treatment enhanced the accumulation of 7-hydroxymethyl chlorophyll
a in the mutant. It might be due to the stimulation of chlorophyll
b to chlorophyll
a conversion by the treatment. These results suggest that chlorophyll degradation products are not properly reduced in this mutant. This mutant will be a clue to understanding the regulation mechanism of chlorophyll degradation.
View full abstract
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Mari Mochimaru, Takashi Maoka, Shinichi Takaichi
Pages
0563
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
FREE ACCESS
The genus
Nostoc is terrestrial, highly drought-tolerant, and has N
2-fixing activity. We previously analyzed carotenoid compositions from the genera
Nostoc and
Anabaena, and identified functions of carotenogenesis genes. There was diversity in carotenoid compositions, and this is due to presence or absence, and substrate specificities of carotenogenesis genes.
In this study, we extracted pigments from
Nostoc commune NIES-24 (=IAM M-13), purified them, and identified each compound with their profiles of C
18-HPLC, MS, NMR, and CD: β-carotene 50% (mol%), β-cryptoxanthin 1%, zeaxanthin 2%, caloxanthin 5%, nostoxanthin 11%, echinenone 20%, canthaxanthin 2%, myxol 2'-fucoside 4%, and 2-OH-myxol 2'-fucoside 5%. In this species, many carotenogenesis genes seem to function, considered from the result so many compounds were obtained. This is the second species to possess 2-OH-myxol 2'-fucoside; the first is
Thermosynechococcus elongatus BP-1. On the other hand,
N. commune had no ketomyxol glycosides, which is observed in some
Nostoc and
Anabaena species.
View full abstract
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Shinichi Takaichi, Mari Mochimaru
Pages
0564
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
FREE ACCESS
Cyanobacteria essentially have various carotenoids for light-harvesting and photoprotection. They have unique carotenoids of carotenoid methylpentosides and ketocarotenoids, such as myxol 2'-fucoside and echinenone. Genome DNA sequences of more than 40 species and/or strains have been determined, but carotenoids in some species are not identified. We performed the BLAST-search for carotenogenesis genes with the chosen query sequences, whose functions have already been confirmed. We found that some homologues have no functions themselves or there are no carotenoids of products. Some genes cannot be found from homologous search but products are present. The different compositions of carotenoids among these species might be due to the presence or absence of certain gene(s), or to different enzyme characteristics including substrate specificities. Two distinct β-carotene hydroxylases, CrtR and CrtG, are bifunctional enzymes. Two distinct β-carotene ketolases, CrtO and CrtW, are properly used in two pathways depending on the species.
View full abstract
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Kazuyuki Watabe, Tadashi Mizoguchi, Tsukasa Hatano, Jiro Harada, Hitos ...
Pages
0565
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
FREE ACCESS
In photosynthesis, primary processes occur in photosystems composed of light-harvesting antenna complexes (LHs) and reaction centers (RCs). Most purple photosynthetic bacteria contain bacteriochlorophyll(BChl)-
a, while some bacteria contain BChl-
b whose Q
y-absorption band is more red-shifted than that of BChl-
a.
In this study, we isolated two types of RCs from
Rhodobacter sphaeroides 2.4.1 containing BChl-
a and
Blastochloris (
Rhodopseudomonas)
viridis DSM133 containing BChl-
b. In order to examine the effect of molecular structures of composite BChls upon the photoreaction efficiency, we reconstructed photoreaction units according to the procedures reported by Miyake and his colleagues: BChl-
a type RCs (λ
max = 867 nm) and BChl-
b type RCs (λ
max = 973 nm) were embedded in L-α-phosphatidylcholine membranes together with either exogenous BChl-
a (λ
max = 771 nm) or Chl-
a (λ
max = 661 nm) as an energy-donating pigment. We measured the light-induced absorbance change and photocurrent response of the reconstructed photoreaction units.
View full abstract
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Mamiko Kita, Ritsuko Fuji, Masahiko Iha, Hideki Hashimoto
Pages
0566
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
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Cladosiphon okamuranus is an original strain of a brown alga in Okinawa and never found in the other place. Recently Iha et al. was successful to culture a discoidal form of the alga, which is a precursor of the alga body, and found that fucoxanthin contents was different in addition to the difference in the total weight when the light color differed during culture. Brown algae contain fucoxanthin-chlorophyll
a/
c proteins (FCP) as antenna complexes. FCP is a membrane protein with molecular weight of approximately 20000 and is spread over the oceanic photosynthetic prokaryote, although it has been isolated from only a few diatoms and brown algae and the 3-dimensional structure is not yet known. Thylakoid content is much higher in the discoidal form than that in the algae body, so there is an advantage of scale. We established the isolation and purification method of the FCP from the discoidal form.
View full abstract
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Tadashi Mizoguchi, Yuki Kimura, Chihiro Nagai, Michio Kunieda, Hitoshi ...
Pages
0567
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
FREE ACCESS
Chlorophylls(Chls)-
c are widely distributed and abundant in chromophyte algae and some prokaryotes together with Chl-
a and carotenoids. The pigments are proved to function in light-absorbing pigments in photosynthesis. However, the detailed molecular structures of Chls-
c have not yet been determined in terms of the conformation of the unique acrylic residue at the 17-position as well as the stereochemistry at the chiral 13
2-position. In this study, Chl-
c1 and Chl-
c2 were efficiently extracted from a diatom
Chaetoseros calcitrans and the former (8-ethyl) and the latter (8-vinyl) were carefully separated using HPLC. The conformation of the acrylate was unambiguously determined to be
cisoid around the C17-C17
1 bond using
1H-
1H NOE correlations. Moreover, we developed a novel chiral HPLC method for quantitative analysis of a pair of enantiomeric isomers at the 13
2-position, which enabled us to determine the stereochemistry to be 13
2-(
R)-configuration in combination with CD spectroscopy.
View full abstract
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Norikazu Ohnishi, Yuichiro Takahashi
Pages
0568
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
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Photosystem I is a chlorophyll protein complex, which is consist of 14 subunits and mediates the electron transfer from plastocyanin to ferredoxin. PsaH is one of the subunits of photosystem I and has been proposed to be required for state transitions. However, the molecular mechanism of its contribution to state transitions has not been clarified. In the present study, we generated knock down transformants of
PsaH by means of RNAi technique from a green alga
Chlamydomonas reinhardtii. After screening of 15 paromomycin-resistant transformants, we obtained 2 lines of transformant in which the accumulation of PsaH protein decreased to approximately 20% of that in wild-type cells. We are now analyzing the state transitions of these transformants by means of low temperature fluorescence spectra and will discuss the role of PsaH on the association of minor LHCII proteins with photosystem I.
View full abstract
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Chihiro Azai, Jiro Harada, Toru Kondo, Shigeru Itoh, Hirozo Oh-oka
Pages
0569
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
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The green sulfur bacteria are strictly anaerobic photo-autotrophic organisms. Their reaction center (RC) complexes are classified into the PS I-type RC. Their RC core protein is a homodimer, while the other well-known ones are heterodimers whose tertiary structures have already been determined. The detailed mechanisms on electron transfer reactions within green sulfur bacterial RC remain to be resolved because it is too unstable to be analyzed by various biochemical and spectroscopic methods under aerobic conditions. Molecular identification of all electron transfer cofactors has not yet been done. Especially, it has still been controversial whether quinone molecule exists and functions as a secondary acceptor. Molecular biological methods, such as the addition of some affinity tag and/or the site directed mutagenesis, are therefore expected to cope with these issues. In this study, we constructed a gene expression system to introduce the tagged core protein in the green sulfur bacterium
Chlorobium tepidum.
View full abstract
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Isamu Ikegami, Yuji Watanabe, Yosuke Morikawa, Sinya Ohtake
Pages
0570
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
FREE ACCESS
Most Chl a in the PS1RC complexes was removed by ether treatment without any loss of P700, yielding the antenna-depleted complexes with a Chl a/P700 ratio of 13. (1) Chl b, Chl a-epimer or Chl d, which have a side chain different from Chl a, could bind to the Chl a-binding sites of the antenna-depleted PSIRC complexes and transfer its excitation energy to P700 with an efficiency as high as Chl a, except Chl d, of which excited-state level is almost equal to that of P700. (2) Metal-substituted Chl a could bind to the Chl a-binding sites; the excitation-energy could be transferred to P700 with Zn-, Cd- or Sn-Chl a, but not with Hg-, Cu- or Fe-Chl a, possibly depending on the lifetime of their excited states. (3) Zn-Chlorophyllide a lacking phytol, could bind twice more efficiently to the complexes, but with less excitation-energy transfer than Zn-Chl a.
View full abstract
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Chihiro Kamidaki, Souji Ishizaka, Tomoyasu Noji, Tsutomu Kajino, Yoshi ...
Pages
0571
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
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Silica mesoporous materials (SMM) have nano-scale pores its inside and adsorb various molecules. We reported the introduction of light-harvesting LH2-membrane protein complex (137 kDa) of photosynthetic purple bacteria into FSM, a type of SMM with rather small inner pores. In this study we introduced the huge trimer of photosystem I (PSI) reaction center complex (1068 kDa), which was purified from a thermophilic cyanobacterium Thermosynechococcus elongatus and has a diameter of 21 nm, into different types of SMM that are different in pore sizes and outer shapes. The PSI adsorbed into these SMM showed high activity to undergo photochemical reaction of chlorophyll and electron transfer, and retained native pigment protein structure as confirmed by the laser spectroscopy. Moreover, adsorption to SMM increased the heat stability of photochemical activity and the protein structure by 10 ~ 15 degree. Most of the SMM-PSI conjugates revealed high activity in the photo-reduction of methyl viologen.
View full abstract
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Hisako Kubota, Azusa Kurihara, Naoki Mizusawa, Hajime Wada
Pages
0572
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
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Recently, X-ray crystallographyic analysis of cyanobacterial photosystem II (PSII) elucidated that not only protein subunits but also many cofactors such as chlorophylls, metals, and lipids are present as structural component in the complex. Lipid molecules might play an important role in PSII complex. Moreover, lipids are used as a substrate for lipid modification of extrinsic proteins, Psb27 and PsbQ of PSII, although they have not seen in the crystal structure. Psb27 and PsbQ are modified with a diacylglycerol and a diacylglycerol and an acyl group, respectively. However, the reasons why these proteins are modified with lipids and they are differently modified with lipids are still unknown. In this study, we generated
Synechocystis sp. PCC 6803 strains, which express chimeric proteins of PsbQ and Psb27 that signal peptides are interchanged each other to change the lipid modification of the proteins.
View full abstract
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Naoki Mizusawa, Shinya Sakata, Isamu Sakurai, Naoki Sato, Hajime Wada
Pages
0573
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
FREE ACCESS
We examined the effect of lack of digalactosyldiacylglycerol (DGDG) on the heat sensitivity to photosynthetic machinery using a
dgdA mutant of
Synechocystis sp. PCC6803 that is defective in the biosynthesis of DGDG. The mutant cells showed normal growth under low light at 30
oC. However, the growth of mutant cells was retarded compared to wild-type cells when the growth temperature was changed to 38
oC. The growth retardation at 38
oC became more apparent under high light. Experiments on photoinhibition of photosynthesis showed that both photodamage and repair processes of photosynthesis were severely affected at 38
oC in the mutant cells compared to the wild-type cells. The oxygen-evolution activity of mutant cells was more easily inactivated by dark-incubation at 38
oC and by incubation with hydroxylamine than that of wild-type cells. These results suggest that the decreased stability of oxygen-evolution system in mutant cells results in the increased sensitivity to light and high temperature stresses.
View full abstract
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Maki Utsumi, Hisako Kubota, Naoki Mizusawa, Hajime Wada
Pages
0574
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
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Photosystem II (PSII) performs important reactions in chloroplasts and cyanobacteria: production of oxygen from light-driven oxidation of water and electron transfer from water to plastoquinone. The precomplex of PSII is transformed into an active complex, which consists of more than 20 kinds of proteins, chlorophylls, metals and plastoquinones. PSII complex is constructed by attachment of the intrinsic proteins CP43/CP47 to core protein D1/D2, incorporation of several small subunits, and assembly of the water-splitting system. Recently the cyanobacterial PSII structure by X-ray crystallography was reported, but the process of how the components are assembled remains unknown. Here we report our study on an extrinsic protein Psb27, which plays an important role in building the water-splitting system. PSII complexes isolated from the strain expressing Histidine-tagged Psb27 were analyzed by Blue-Native PAGE and 2D SDS-PAGE, and the obtained results indicated that Psb27 binds to monomer complexes, which have different subunit compositions.
View full abstract
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Shinya Sakata, Hisako Kubota, Isamu Sakurai, Naoki Mizusawa, Hajime Wa ...
Pages
0575
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
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Recently, it have been demonstrated that several proteins are involved in the assembly of the Photosystem II (PSII) complex. One of these proteins, Psb28, which is conserved from cyanobacteria to higher plants, predicted to be an extrinsic protein bound to cytosolic side of the PSII complex.
In this study, we constructed
psb28-deletion mutant of
Synechocystis sp. PCC 6803 to elucidate the role of the Psb28 protein. The mutant showed normal photoautotrophic growth and oxygen-evolving activity under normal growth conditions. Analysis of the PSII complex purified from the strain expressing a His-tagged Psb28 protein revealed that Psb28 is mainly associated with assembly intermediates of PSII lacking CP43 which binds antenna chlorophylls.
Based on these observations, the role of Psb28 in the assembly of the PSII complex is discussed.
View full abstract
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Mahbuba Khatoon, Kayo Inagawa, Miho Yoshioka, Noriko Morita, Yasusi Ya ...
Pages
0576
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
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Light and heat stresses induce damage to Photosystem II (PSII). We investigated relationship between the stress-induced damage to the D1 protein and unstacking of spinach thylakoids. For efficient migration of the damaged PSII complexes from the grana to the stroma thylakoids where the damaged PSII is repaired, the grana should become unstacked. We used digitonin fractionation method to estimate membrane unstacking. During strong illumination light intensity 1,000 μmol photons m-2 s-1 of PSII at 25degree, the thylakoids became markedly unstacked. Heat stress (40 degree) in the dark also induced thylakoid unstacking. Interestingly, the extent of D1 degradation was proportional to the rate of the thylakoid unstacking. By contrast, the thylakoid unstacking did not take place at 4 degree under high light and the D1 degradation was prevented. These results suggest that the thylakoid unstacking is necessary for efficient degradation of the damaged D1 protein.
View full abstract
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Miho Yoshioka, Keisuke Komayama, Kayo Inagawa, Yasusi Yamamoto
Pages
0577
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
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It has been suggested that FtsH proteases are abundant in the stroma thylakoids and the grana margin because these domains are the site of repair of the damaged D1 protein in the thylakoids. In the present study we assayed the relative amount of the protease in the thylakoids, grana, stroma thylakoids, PSII membranes and PSII core. The active FtsH protease forms hexameric ring structure, but it is not clear whether the hexameric proteases are present in every membrane domain (fraction). We analyzed the structure of the FtsH proteases under control dark conditions, as well as heat or light stress conditions, using clear native gel electrophoresis. This work was supported by the JSPS fellowship (DC2).
View full abstract
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Masayuki Komura, Hiroyuki Miyake, Atsushi Yamagishi, Yutaka Shibata, I ...
Pages
0578
Published: 2009
Released on J-STAGE: October 23, 2009
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Lichens are photoautotrophic symbionts of fungi and algae and survive even under the drought environments. It has been suggested that dried lichens immediately convert excess light energy into heat to prevent the photoinhibition in photosystem II (PSII). A recent report suggests that the chlorophyll emitting fluorescence at 740 nm (Chl740) is present and acts as an energy quencher in PSII in lichen [1]. We used the same type of lichens and measured ultrafast fluorescence decay in dry lichen cells by the combination of upconversion and streakcamera system at 4-300 K to study whether and where Chl740 is present in PSII.
The fluorescence of Chl740 rising within 1 ps after laser excitation, decaying in several picoseconds was observed. The rate of EET and quenching process in PSII was estimated.
We also discuss with respect to other two different types of lichens.
[1] Veerman et al, Plant Physiol., 145, 997-1005 (2007)
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Yuichiro Shimada, Hiroyuki Suzuki, Tohru Tsuchiya, Tatsuya Tomo, Takum ...
Pages
0579
Published: 2009
Released on J-STAGE: October 23, 2009
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Photosynthetic water oxidation is performed by the light-driven S-state cycle in the oxygen-evolving complex of photosystem II. While ligation to the Mn cluster from several amino acid residues has been proposed by X-ray crystallographic studies, details of the reaction processes involving these residues are not clarified. In this study, we constructed a mutant of
Synechocystis sp. PCC 6803 by substituting glutamine for glutamate at the position 354 of CP43 (CP43-Glu354), and investigated the oxygen-evolution processes using spectroscopic measurements including FTIR analysis. The oxygen-evolving activity of the mutant was approximately 20% of wildtype. The oscillation pattern of flash-dependent delayed luminescence suggested that the transition beyond the S
3 state was blocked in the mutant. FTIR difference spectroscopy revealed that CP43-Glu354 functioned as a ligand to the Mn cluster, and changed its coordination structure in the S
1-to-S
2 transition. Based on these findings, we discuss the reaction processes of oxygen evolution involving CP43-Glu354.
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Tatsuya Tomo, Seiji Akimoto, Tohru Tsuchiya, Michitaka Fukuya, Kazunor ...
Pages
0580
Published: 2009
Released on J-STAGE: October 23, 2009
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We isolated highly-purified photosystem (PS) II reaction center (RC) complexes from the cyanobacterium
Synechocystis sp. PCC 6803 using a histidine tag introduced to the 47 kDa chlorophyll protein, and characterized their spectroscopic properties. Purification was carried out in a one-step procedure after isolation of PS II core complex. The polypeptide composition and pigment stoichiometry were the same as in spinach. Overall absorption and fluorescence properties were very similar to those of spinach PS II RCs. However, a clear band-shift of pheophytin
a and β-carotene was observed. Reasons for these differences, and RC composition, are discussed on the basis of the three-dimensional structure of complexes.
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Tomoyasu Noji, Chihiro Kamidaki, Keisuke Kawakami, Jian-Ren Shen, Tsut ...
Pages
0581
Published: 2009
Released on J-STAGE: October 23, 2009
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Silica mesoporous materials has nano size pore in large quantity, and are an inorganic materials. We so far succeeded in introducing some membrane proteins of photosynthesis into silica mesoporous materials. LHII isolated from thermophilic purple photosynthetic bacterium
T.tepidum and PSI from
T.elongatus maintained activity and increased thermal stability after introduced into silica mesoporous materials.
We introduced the photosynthetic photosystem II (PSII) pigment-protein complex into silica mesoporous materials. The dimer complex of PSII reaction centers (756 kDa) were purified from a thermophilic cyanobacterium
Thermosynechococcus vulcanus. The adsorption to silica mesoporous materials partially decreased the oxygen evolution activity without significant effects on the pigment assembly and the protein structure based on the absorption and fluorescence spectrum. The high thermal stability of oxygen evolution reaction was maintained inside silica. We study the effects of pore size too.
PSII adsorbed to silica can be used as a new material for the artificial reactions.
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Bjorn Lundin
Pages
0582
Published: 2009
Released on J-STAGE: October 23, 2009
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ATP is a nucleotide and the major energy currency, but there are also other nucleotides such as AMP, ADP, GMP, GDP and GTP. Chloroplast metabolism has mostly been associated with ATP, but GTP has been shown to have a role in integration of light harversting complexes into the thylakoid. Here, we have demonstrated the occurrence of nucleotide-dependent processes in the lumenal space of spinach by bringing evidence first for nucleotide (ATP) transport across the thylakoid membrane, second for nucleotide inter-conversion (ATP to GTP) by a nucleoside diphosphate kinase, and third the discovery that the PsbO extrinsic subunit of PSII complex can bind and hydrolyse GTP to GDP.
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Chihiro Watanabe, Takushi Hachiya, Kentaro Takahara, Maki Kawai, Hirof ...
Pages
0583
Published: 2009
Released on J-STAGE: October 23, 2009
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In the plant respiratory-chain, the ATP-uncoupling cyanide-resistant pathway, catalyzed by alternative oxidase (AOX), is supposed to prevent ROS production and maintain carbon to nitrogen (CN) ratio. In this study, we investigated whether AOX contributes to control CN ratio under low-N condition. We treated
Arabidopsis thaliana WT and
aox knock-out transgenic with short- or long-term low-N. In WT, cyanide-resistant respiratory rate and
AOX expression increased after the short-term low-N treatment. Total respiratory rate in
aox was similar to that of WT. Some intermediates increased under low-N condition, and those of
aox were lower than WT. In long-term low-N treatment, growth of
aox was greater than that of WT. These results suggest that the decrease of intermediate levels in
aox leads to higher growth in long-term N deficiency. There was no significant difference in CN ratio between the genotypes, suggesting that AOX does not contribute to control CN ratio under low-N condition.
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Sakiko Nagashima, Keizo Shimada, Kenji Nagashima
Pages
0584
Published: 2009
Released on J-STAGE: October 23, 2009
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A photoheterotrophic bacterium,
Rubrivivax gelatinosus, possesses HiPIP, high-potential (HP), low-potential (LP) cytochrome
c8 and cytochrome
c4 which can work as electron donors to the photosynthetic reaction center. Additionally, we have recently identified an isotype of HP cytochrome
c8 from a mutant strain that spontaneously suppressed previous mutant deficiencies in the photosynthetic capabilities of the four other electron carriers. In this study, we demonstrated that this iso-HP cytochrome
c8 potentially supports cyclic electron transfer during photosynthetic growth. Furthermore, a 10-kb DNA region around the gene for the iso-HP cytochrome
c8 was determined. Within this region we found a gene cluster (
nirESMCFDLGHJN) for cytochrome
cd1-type nitrite reductase (Nir). The iso-HP cytochrome
c8 gene is 59% homologous to NirM from
Pseudomonas aeruginosa, which donates an electron to Nir. In order to define physiological roles of this cytochrome in
Rvi. gelatinosus, the construction of a mutant devoid of iso-HP cytochrome
c8 is in progress.
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Yuji Suzuki, Takeaki Miyamoto, Ryuichi Yoshizawa, Tadahiko Mae, Amane ...
Pages
0585
Published: 2009
Released on J-STAGE: October 23, 2009
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Effects of an overexpression of
RBCS on Rubisco content, photosynthesis and plant growth were examined in rice (
Oryza sativa L.). Rubisco content in the transformant was significantly greater in the uppermost, fully expanded leaves, but decreased to levels similar to those in wild-type plants in the lower leaves. Although the activation state of Rubisco was lower in the uppermost, fully expanded leaves of the transformant, it recovered to its full level in the lower leaves. As a result, the photosynthetic rate did not differ in leaves at the same position between these genotypes. Similarly, whole plant biomass did not differ between these genotypes. Thus, we conclude that although the overexpression of
RBCS led to an enhancement of Rubisco protein content in the uppermost, fully expanded leaves, it does not result in increased photosynthetic rates or plant biomass, due to an apparent down-regulation in its activation state.
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Masaaki Tachibana, Sae Kikutani, Megumi Fujii, Yusuke Matsuda
Pages
0586
Published: 2009
Released on J-STAGE: October 23, 2009
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Marin diatoms can concentrate inorganic carbon (Ci) under air level CO
2 pressure. Accumulated Ci is thought to be delivered to Rubisco efficiently via aids of Carbonic anhydrases (CAs). In the marine diatom
Phaeodactylum tricornutum, two chloroplastic CAs, PtCA1 and PtCA2, were identified which form cluster on the girdle lamella. At least 8 putative CA genes are found in
P. tricornutum genome. In the present study, these genes were cloned. Expressions of these CA genes were analyzed by RT-PCR. These putative CA genes were also fused with
egfp. As a result, majority of these putative CAs were localized within the four-layered membrane systems of the chloroplast. Function of these putative CAs will be discussed.
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Yukiko Yamazaki, Yusuke Matsuda
Pages
0587
Published: 2009
Released on J-STAGE: October 23, 2009
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The Β-type carbonic anhydrase (PtCA1) localizes as clustered particles on the girdle lamellae in the chloroplast of the marine diatom
Phaeodactylum tricornutum . PtCA1 has been suggested to play an important role in CCM but mechanisms and physiological functions of PtCA1 are yet to be elucidated. PtCA1 forms cluster via a function of the amphipathetic helix at its C-terminus which occurs only in the chloroplast. This indicates that particle formation requires some stromal factors. As the first candidate, Rubisco was localized using GFP labeling but results unlikely co-localize with PtCA1. Immnopurecipitation assay revealed that PtCA1 antibody pull down only PtCA1 and its paralogue PtCA2. These results suggest that stromal factors mightn't be proteins or that interactions of particle components are fairly weak which doesn't fully function outside the stroma. The activity of recombinant PtCA1 was measured under conditions mimicking the stroma.
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Hirobumi Nakano, Yosuke Yamahara, Shin-ichiro Ozawa, Yuichiro Takahash ...
Pages
0588
Published: 2009
Released on J-STAGE: October 23, 2009
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Aquatic photosynthetic organisms can acclimate to CO
2-limiting stress conditions by inducing the carbon-concentrating mechanism (CCM). It is not known how cells can sense the decrease of environmental CO
2 concentration leading to induce CCM. We focus on the CCM1-regulatory protein complex found in
Chlamydomonas reinhardtii (Kohinata
et al. 2008). From a transgenic cells harboring tagged CCM1 gene, we have isolated the CCM1-complex by affinity chromatography followed by MS analyses and identified a component of the complex. Here we will present a series of data concerning to a newly identified protein P80, which is one of the component of the CCM1 complex in
Chlamydomonas cells.
1) Kohinata
et al. Plant Cell Physiol. 49:273-283 (2008)
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Aki Kato, Masahiro Inouhe
Pages
0589
Published: 2009
Released on J-STAGE: October 23, 2009
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Plant cells live and grow in a manner totally dependent upon sugars. Externally applied sugars support the long-term growth of cells in the tissue and/or cell cultures of various plant species. However, mannose (Man) is not usable as external carbon source by many plants. We compared soluble sugar compositions and metabolisms in various types of azuki-bean cells cultivated in the presence or absence of Man. We found that the heterotrophic cells commonly required Suc and wall polysaccharide syntheses via sugar-phosphates and sugar-nucleotides pathways for their growth activities. Also the levels of sugar-phosphates and the isomerase activities appeared important for cell ability to use uncommon sugars as external carbon sources. Callus cells can utilize Man as a sole carbon-source on agar media but not in liquid cell-suspension culture, suggesting an important timing or balance between induction of PMI enzyme and temporal accumulation of M-6-P in cells.
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Tadamasa Sasaki, Haruka Takeda, Akina Tsunoda, Shigetoshi Miura, Hiroa ...
Pages
0590
Published: 2009
Released on J-STAGE: October 23, 2009
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Sucrose synthase catalyzes a first step for sucrose metabolism at sink organs. The sucrose catabolites are utilized for cellulose and starch synthesis, therefore sucrose synthase is one of the key enzyme for storage material synthesis in sink organs. In rice, three isozymes of sucrose synthase have been identified (
Susy1,
Susy2 and
Susy3). We have found the involvement of SPK (Seed-development specific Protein Kinase) for the phosphorylation of Susy in rice pre-mature seeds. Thus we investigated the effect for phosphorylation of Susy by SPK, and we obtained data where it was indicated that the rice sucrose synthase were regulated by phosphorylation in vitro. Further more, we revealed that rice Susys could utilize ADP as well as UTP, in both reactions, sucrose degradation and sucrose synthesis, respectively.
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Tomoki Tabuchi, Masayo Demuratani, Kumi Otori, Noriaki Tanabe, Masahir ...
Pages
0591
Published: 2009
Released on J-STAGE: October 23, 2009
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We isolated an Arabidopsis mutant (
sicy-192) whose cotyledon greening was inhibited by treatments with sugars, such as sucrose and glucose. In the mutant, the gene encoding plastidic alkaline/neutral invertase (INV-E) was point-mutated at codon 294, with Tyr substituted for Cys. The greening of cotyledons in the knockout-INV-E lines was not inhibited by treatment with the sugars. In addition, the knockout-INV-E lines expressing an
INV-E:C294Y gene had the same phenotype as
sicy-192. On treatment with sucrose, the expression of photosynthesis-related genes was weaker in seedlings of mutant plants than wild-type seedlings, while the activity of nitrate reductase was stronger in the mutant plants than wild-type plants. These findings suggest that INV-E is regulated for maintenance of the level of sucrose or its metabolites in plastids, preventing an imbalance between the supply of nitrogen to cells via nitrate assimilation and the demand for nitrogen for development of the photosynthetic apparatus.
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Kentarou Kaneko, Chie Yamada, Ai Yanagida, Aya Kitajima, Kimiko Itoh, ...
Pages
0592
Published: 2009
Released on J-STAGE: October 23, 2009
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We investigate the function of nucleotide pyrophosphatase/phosphodiesterase (NPP) in rice. Six NPP isozyme genes have been found in rice. Previously, we purified and characterized NPP1, 2 and 6. NPP1 and 6 were divided into ADP-glucose hydrolyzing type, while NPP2 was nucleotide hydrolyzing type. All NPP isozymes were conjugated with Con A-recognized saccharide chains, and existed as 70-74 kDa homooligomeric forms. Furthermore, the expression and targeting of
NPP-GFP fusion genes revealed that NPPs localize within the chloroplasts. In the present study, we analyzed phenotypic divergence, starch and free sugar contents in
NPP1 overexpressed and knocked-out mutant plants. The former plants exhibited growth stimulation of young roots, decrease of starch accumulation and increase of free sugar contents. Intriguingly, the weights of grains harvested from
npp1 or
npp6 mutants were significantly increased. The overall results strongly suggested that ADP-glucose hydrolyzing NPPs are involved in controlling root elongation through regulating the starch synthesis.
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Michiko Shiga, Takuro Ogawa, Daisuke Seo, Yuko Nishimoto, Hidehiro Sak ...
Pages
0593
Published: 2009
Released on J-STAGE: October 23, 2009
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Anaerobic photosynthetic green sulfur bacteria utilize sulfur compounds such as sulfide and thiosulfate as electron donors of photosynthesis. Previously, we reported purification of four proteins (SoxYZ, SoxB, SoxAX-CT1020, SoxJ) involved in thiosulfate oxidation from
Chlorobaculum tepidum cells. Some thiosulfate oxidizing bacteria have
CT1020 homolog, named
soxK, but its function was unknown. We have over-expressed SoxA, SoxX and SoxK in
E. coli, and found that SoxK strengthens the binding of SoxA and SoxK, and stimulates the activity. Accordingly, SoxK is designated as SAXB (SoxAB-binding protein). From comparative genome analyses, about one third of thiosulfate bacteria have
soxK homologues, and the deduced SoxA and SoxX proteins cluster together according to the presence and absence of
soxK. Flavoprotein SoxJ is a homologue of flavoprotein subunit of flavocytochrome
c. SoxJ itself has no thiosulfate oxidizing activity, but it accelerated the reaction rate about twice when added to the mixture of SoxAXK, SoxB and SoxYZ.
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Naomi Hosoya-Matsuda, Junya Kmijou, Toru Hisabori, Kazuhito Inoue
Pages
0594
Published: 2009
Released on J-STAGE: October 23, 2009
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Thioredoxin is a small ubiquitous protein that is involved in the dithiol-disulfide exchange reaction, by way of two cysteine residues located on the molecule surface. In order to elucidate the role of thioredoxin in
Chlorobaculum tepidum, an anaerobic green sulfur bacterium that uses various inorganic sulfur compounds as an electron donor under strict anaerobic conditions for growth, we applied the thioredoxin affinity chromatography method. In this study, 37 cytoplasmic proteins were captured as thioredoxin target candidates, including proteins involved in sulfur assimilation. Furthermore, six of the candidate proteins were members of the reductive tricarboxylic acid cycle. The redox sensitivity of three of them was examined: citrate lyase, citrate synthase, and malate dehydrogenase, using their recombinant proteins. Based on the information relating to the target proteins, the significance of thioredoxin as a reductant for the metabolic pathway in the anaerobic photosynthetic bacteria is discussed.
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Wanwipa Ittarat, Takuro Ogawa, Daisuke Seo, Kazuyoshi Ueda, Michiko Sh ...
Pages
0595
Published: 2009
Released on J-STAGE: October 23, 2009
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We have prepared 4Fe-4S type ferredoxin (Fd) from the cell extract of
H. modesticaldum by ammonium sulfate fractionation and anion-exchange chromatography. Crude RC-rich fractions were prepared under anaerobic conditions by solubilization with detergents of the membranes of the bacterium, and one of them showed NADP
+ photoreduction activity in the presence of Fd I from
H. modesticaldum and Fd-NADP
+ reductase (FNR) from spinach. The active fractions partially purified by sucrose-density gradient ultracentrifugation reduced NADP
+ at 12 μmole μmole BChl
-1 hr
-1. The peptides in the active fractions were analyzed by SDS-PAGE and TOF-Mass spectrometer. Although the whole genome sequence of this bacterium has been elucidated, the gene encoding FNR has not yet been identified. From the cell extract, we obtained several partially purified fractions containing NADPH-DPIP diaphorase activity, but the major active fraction has no FNR activity.
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Mitsunori Katayama
Pages
0596
Published: 2009
Released on J-STAGE: October 23, 2009
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Phototropism is a growth movement towards a source of light. It is observed among eukaryotes such as plant, alga, fungi, and also among some prokaryotes. Last year, we reported that several species of cyanobacteria belong to genera
Calothrix and
Scytonema (both are grouped into subsection IV) exhibit a growth response that corresponds to photropism.
Here we report that
Fischerella sp. that is grouped into subsection V exhibits phototropism. Subclass V cyanobacteria have filament of cells as a unit and their cells divide more than one plane. The
Fischerella sp. isolated in this study shares some characteristics with other genera of cyanobacteria. That is, tapered filament, living on land, growing towards blue light. Likewise
Calothrix, Fischerella includes species that does and does not exhibit phototropism. Phylogenetic relationship of cyanobacteria that exhibit photropism is to be further investigated.
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Jun Kaseda, Daiji Gojima, Kazuhiro Nagahama, Masayosi Matsuoka
Pages
0597
Published: 2009
Released on J-STAGE: October 23, 2009
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Genetically amenable cyanobacteria are most suited for analyzing the mechanism of plant-type oxygen evolution. We have constructed cyanobacterial strains of
Synechococcus elongatus PCC 7942 which carried thermostable D1/D2 heterodimer of photosystemII(PSII) from
Thermosynechococcus vulcanus. In this study, a Histag was inserted at CP47 subunit of PSII, enabling purification of PSII via affinity chromatography. For this purpose,
rps12 -mediated gene replacement scheme was employed to construct recombinant strains carrying
psbB gene with Histag sequence at the 3' end. PCR analysis of the recombinant strains confirmed that chromosomal
psbB-psbT operon had been replaced with
psbB-histag::[rps12-kan]-psbT.
To circumvent the adverse effect on the downstream
psbT gene, a surrogate
psbB promoter was inserted upstream the
psbT gene to make a recombinant strain GRPS801.These strains as well as strains devoid of [
rps12-kan ] cassette are now tested for one-step purification of PSII complex.
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Yuki Wakamatsu, Takuro Ogawa, Mari Kobayashi, Rei Narikawa, Kazuhito I ...
Pages
0598
Published: 2009
Released on J-STAGE: October 23, 2009
CONFERENCE PROCEEDINGS
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Oxidative stress and its protection are critical for survival of oxygenic phototrophs. We have demonstrated that superoxide stress induced expression of iron-sulfur center assembly genes in the sufB operon via a putative transcriptional repressor sll0088(SufR), which possesses an iron-sulfur center motif. Here we studied the SufR homolog protein of
Thermosynechococcus elongatus BP-1(TeSufR), which was expressed in
E. coli . When TeSufR was prepared anaerobically, it showed [4Fe-4S]-type EPR signals. When TeSufR was prepared aerobically, it showed a [1Fe-0S]-type EPR signal. We also reconstituted the TeSufR apoprotein
in vitro with iron under reducing conditions. We obtained similarly [4Fe-4S]-type TeSufR under anaerobic conditions, and [1Fe-0S]-type TeSufR under aerobic conditions. Further exposure of TeSufR to the aerobic solution revealed that [4Fe-4S]-type is more labile than [1Fe-0S]-type. Possible role and its physiological relevance of these two iron-sulfur centers will be discussed.
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Soichirou Satoh, Tohru Tsuchiya, Mamoru Mimuro
Pages
0599
Published: 2009
Released on J-STAGE: October 23, 2009
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Gloeobacter violaceus PCC 7421 is a thylakoid-lacking unicellular cyanobacterium. In its complete genome sequence, however, we found a gene (
glr0898) homologous to
vipp1 (vesicle-inducing protein in plastids 1) that is known to be essential for thylakoid membrane formation in other cyanobacteria and chloroplasts [1]. Amino acid sequence of Glr0898 was also similar to that of phage shock protein A (PspA) that is not involved in thylakoid membrane formation. To investigate the function of
glr0898 gene
in vivo, we introduced a
glr0898 gene into
Synechocystis sp. PCC 6803 cells, and subsequently knocked-out the endogenous
vipp1 genes of the
glr0898-expressing transformants. These transformant cells showed that growth rates, absorption spectra and chlorophyll contents were nearly identical to those of wild type cells. These results suggested that the
glr0898 complemented the loss of
Synechocystis vipp1 in the transformant cells.
[1] Westphal et al. (2001)
Proc. Natl. Acad. Sci. USA, 98, 4243.
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Hidehisa Yoshimura, Masayuki Ohmori, Masahiko Ikeuchi
Pages
0600
Published: 2009
Released on J-STAGE: October 23, 2009
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Cyanobacteria are Gram-negative bacteria possessing the plasma membrane and the outer membrane. Cyanobacteria produce extracellular polysaccharides (EPS). It is considered that the EPS protects the cell from environmental stresses. The EPS is generally categorized into three types: lipopolysaccharides (LPS); capsular polysaccharides (CPS); and the released polysaccharides (RPS). To examine the relation between cell-surface structure and stress-defense mechanism in cyanobacteria, the gene which is involved in the Lipid-A biosynthesis of the outer membrane was disrupted in the filamentous cyanobacterium Anabaena sp. strain PCC 7120. The disruptant cells were more sensitive to various stresses than wild-type cells. The amounts of CPS and RPS of the disruptant cells were decreased in comparison with those of wild-type cells. We will discuss the correlation among the outer membrane, the EPS, and the stress-defense mechanism in cyanobacteria.
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