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1.
Photosynth Res ; 154(3): 277-289, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-35976595

RESUMO

This study aimed to clarify (1) which pigment in a photosystem II (PSII) core complex is responsible for the 695-nm emission at 77 K and (2) the molecular basis for the oxidation-induced fluorescence quenching in PSII. Picosecond time-resolved fluorescence dynamics was compared between the dimeric and monomeric PSII with and without addition of an oxidant. The results indicated that the excitation-energy flow to the 695-nm-emitting chlorophyll (Chl) at 36 K and 77 K was hindered upon monomerization, clearly demonstrating significant exciton migration from the Chls on one monomer to the 695-nm-emitting pigment on the adjacent monomer. Oxidation of the redox-active Chl, which is named ChlZ caused almost equal quenching of the 684-nm and 695-nm emission bands in the dimer, and lower quenching of the 695-nm band in the monomer. These results suggested two possible scenarios responsible for the 695-nm emission band: (A) Chl11-13 pair and the oxidized ChlZD1 work as the 695-nm emitting Chl and the quenching site, respectively, and (B) Chl29 and the oxidized ChlZD2 work as the 695-nm emitting Chl and the quenching site, respectively.


Assuntos
Clorofila , Complexo de Proteína do Fotossistema II , Complexo de Proteína do Fotossistema II/metabolismo , Espectrometria de Fluorescência , Oxirredução , Complexos de Proteínas Captadores de Luz
2.
Photosynth Res ; 135(1-3): 285-298, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29151177

RESUMO

Some mosses stay green and survive long even under desiccation. Dissipation mechanisms of excess excitation energy were studied in two drought-tolerant moss species adapted to contrasting niches: shade-adapted Rhytidiadelphus squarrosus and sun-adapted Rhytidium rugosum in the same family. (1) Under wet conditions, a light-induced nonphotochemical quenching (NPQ) mechanism decreased the yield of photosystem II (PSII) fluorescence in both species. The NPQ extent saturated at a lower illumination intensity in R. squarrosus, suggesting a larger PSII antenna size. (2) Desiccation reduced the fluorescence intensities giving significantly lower F 0 levels and shortened the overall fluorescence lifetimes in both R. squarrosus and R. rugosum, at room temperature. (3) At 77 K, desiccation strongly reduced the PSII fluorescence intensity. This reduction was smaller in R. squarrosus than in R. rugosum. (4) Global and target analysis indicated two different mechanisms of energy dissipation in PSII under desiccation: the energy dissipation to a desiccation-formed strong fluorescence quencher in the PSII core in sun-adapted R. rugosum (type-A quenching) and (5) the moderate energy dissipation in the light-harvesting complex/PSII in shade-adapted R. squarrosus (type-B quenching). The two mechanisms are consistent with the different ecological niches of the two mosses.


Assuntos
Adaptação Fisiológica , Briófitas/fisiologia , Briófitas/efeitos da radiação , Secas , Luz Solar , Adaptação Fisiológica/efeitos dos fármacos , Adaptação Fisiológica/efeitos da radiação , Briófitas/efeitos dos fármacos , Dióxido de Carbono/farmacologia , Desidratação , Dessecação , Cinética , Espectrometria de Fluorescência , Temperatura , Fatores de Tempo
3.
Bioconjug Chem ; 27(10): 2469-2479, 2016 10 19.
Artigo em Inglês | MEDLINE | ID: mdl-27571354

RESUMO

The development of additional extraction surfactants for membrane proteins is necessary for membrane protein research, since optimal combinations for the successful extraction of target membrane proteins from biological membranes that minimize protein denaturation are hard to predict. In particular, those that have a unique basal molecular framework are quite attractive and highly desired in this research field. In this study, we successfully constructed a new extraction surfactant for membrane proteins, NPDGC12KK, from the peptide-gemini-surfactant (PG-surfactant) molecular framework. The PG-surfactant is a U-shaped lipopeptide scaffold, consisting of a short linker peptide (-X-) between two long alkyl-chain-modified Cys residues and a peripheral peptide (Y-) at the N-terminal side of long alkyl-chain-modified Cys residues. Using photosystem I (PSI) and photosystem II (PSII) derived from Thermosynecoccus vulcanus as representative membrane proteins, we evaluated whether NPDGC12KK could solubilize membrane proteins while maintaining structure and functions. Neither the membrane integral domain nor the cytoplasmic domain of PSI and PSII suffered any damage upon the use of NPDGC12KK based on detailed photophysical measurements. Using thylakoid membranes of T. vulcanus as a representative biological membrane sample, we performed experiments to extract membrane proteins, such as PSI and PSII. Based on the extraction efficiency and maintenance of protein supramolecular structure established using clear native-PAGE analyses, we proved that NPDGC12KK functions as a novel class of peptide-containing extraction surfactants for membrane proteins.


Assuntos
Proteínas de Membrana/química , Proteínas de Membrana/isolamento & purificação , Tensoativos/química , Fracionamento Químico/métodos , Cisteína/química , Lipopeptídeos/química , Micelas , Peptídeos/química , Complexo de Proteína do Fotossistema I/química , Complexo de Proteína do Fotossistema II/química , Engenharia de Proteínas/métodos , Espectrometria de Fluorescência , Synechocystis/química , Tilacoides/química
4.
Langmuir ; 32(31): 7796-805, 2016 08 09.
Artigo em Inglês | MEDLINE | ID: mdl-27400072

RESUMO

The development of artificial photosynthesis has focused on the efficient coupling of reaction at photoanode and cathode, wherein the production of hydrogen (or energy carriers) is coupled to the electrons derived from water-splitting reactions. The natural photosystem II (PSII) complex splits water efficiently using light energy. The PSII complex is a large pigment-protein complex (20 nm in diameter) containing a manganese cluster. A new photoanodic device was constructed incorporating stable PSII purified from a cyanobacterium Thermosynechococcus vulcanus through immobilization within 20 or 50 nm nanopores contained in porous glass plates (PGPs). PSII in the nanopores retained its native structure and high photoinduced water splitting activity. The photocatalytic rate (turnover frequency) of PSII in PGP was enhanced 11-fold compared to that in solution, yielding a rate of 50-300 mol e(-)/(mol PSII·s) with 2,6-dichloroindophenol (DCIP) as an electron acceptor. The PGP system realized high local concentrations of PSII and DCIP to enhance the collisional reactions in nanotubes with low disturbance of light penetration. The system allows direct visualization/determination of the reaction inside the nanotubes, which contributes to optimize the local reaction condition. The PSII/PGP device will substantively contribute to the construction of artificial photosynthesis using water as the ultimate electron source.


Assuntos
2,6-Dicloroindofenol/química , Proteínas de Bactérias/química , Cianobactérias/enzimologia , Vidro/química , Nanoporos , Oxigênio/química , Complexo de Proteína do Fotossistema II/química , Porosidade
5.
J Am Chem Soc ; 137(40): 13121-9, 2015 Oct 14.
Artigo em Inglês | MEDLINE | ID: mdl-26403467

RESUMO

Introducing appropriate artificial components into natural biological systems could enrich the original functionality. To expand the available wavelength range of photosynthetic bacterial light-harvesting complex 2 (LH2 from Rhodopseudomonas acidophila 10050), artificial fluorescent dye (Alexa Fluor 647: A647) was covalently attached to N- and C-terminal Lys residues in LH2 α-polypeptides with a molar ratio of A647/LH2 ≃ 9/1. Fluorescence and transient absorption spectroscopies revealed that intracomplex energy transfer from A647 to intrinsic chromophores of LH2 (B850) occurs in a multiexponential manner, with time constants varying from 440 fs to 23 ps through direct and B800-mediated indirect pathways. Kinetic analyses suggested that B800 chromophores mediate faster energy transfer, and the mechanism was interpretable in terms of Förster theory. This study demonstrates that a simple attachment of external chromophores with a flexible linkage can enhance the light harvesting activity of LH2 without affecting inherent functions of energy transfer, and can achieve energy transfer in the subpicosecond range. Addition of external chromophores, thus, represents a useful methodology for construction of advanced hybrid light-harvesting systems that afford solar energy in the broad spectrum.


Assuntos
Complexos de Proteínas Captadores de Luz/química , Complexo de Proteína do Fotossistema II/química , Sequência de Aminoácidos , Transferência Ressonante de Energia de Fluorescência , Dados de Sequência Molecular , Espectrometria de Fluorescência
6.
Plant Cell Physiol ; 56(10): 2024-34, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26320210

RESUMO

Cells of a unicellular cyanobacterium strain KC1, which were collected from Japanese fresh water Lake Biwa, formed chlorophyll (Chl) f at 6.7%, Chl a' at 2.0% and pheophytin a at 0.96% with respect to Chl a after growth under 740 nm light. The far-red-acclimated cells (Fr cells) formed extra absorption bands of Chl f at 715 nm in addition to the major Chl a band. Fluorescence lifetimes were measured. The 405-nm laser flash, which excites mainly Chl a in photosystem I (PSI), induced a fast energy transfer to multiple fluorescence bands at 720-760 and 805 nm of Chl f at 77 K in Fr cells with almost no PSI-red-Chl a band. The 630-nm laser flash, which mainly excited photosystem II (PSII) through phycocyanin, revealed fast energy transfer to another set of Chl f bands at 720-770 and 810 nm as well as to the 694-nm Chl a fluorescence band. The 694-nm band did not transfer excitation energy to Chl f. Therefore, Chl a in PSI, and phycocyanin in PSII of Fr cells transferred excitation energy to different sets of Chl f molecules. Multiple Chl f forms, thus, seem to work as the far-red antenna both in PSI and PSII. A variety of cyanobacterial species, phylogenically distant from each other, seems to use a Chl f antenna in far-red environments, such as under dense biomats, in colonies, or under far-red LED light.


Assuntos
Clorofila/análogos & derivados , Cianobactérias/metabolismo , Complexo de Proteína do Fotossistema I/metabolismo , Complexo de Proteína do Fotossistema II/metabolismo , Clorofila/metabolismo
7.
Biochim Biophys Acta ; 1827(4): 529-39, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23416844

RESUMO

Intact fucoxanthin (Fucox)-chlorophyll (Chl)-binding protein I-photosystem I supercomplexes (FCPI-PSIs) were prepared by a newly developed simple fast procedure from centric diatoms Chaetoceros gracilis and Thalassiosira pseudonana to study the mechanism of their efficient solar energy accumulation. FCPI-PSI purified from C. gracilis contained 252 Chl a, 23 Chl c, 56 Fucox, 34 diadinoxanthin+diatoxanthin, 1 violaxanthin, 21 ß-carotene, and 2 menaquinone-4 per P700. The complex showed a high electron transfer activity at 185,000µmolmg Chl a(-1)·h(-1) to reduce methyl viologen from added cytochrome c6. We identified 14 and 21 FCP proteins in FCPI-PSI of C. gracilis and T. pseudonana, respectively, determined by N-terminal and internal amino acid sequences and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analyses. PsaO and a red lineage Chla/b-binding-like protein (RedCAP), Thaps3:270215, were also identified. Severe detergent treatment of FCPI-PSI released FCPI-1 first, leaving the FCPI-2-PSI-core complex. FCPI-1 contained more Chl c and showed Chl a fluorescence at a shorter wavelength than FCPI-2, suggesting an excitation-energy transfer from FCPI-1 to FCPI-2 and then to the PSI core. Fluorescence emission spectra at 17K in FCPI-2 varied depending on the excitation wavelength, suggesting two independent energy transfer routes. We formulated a model of FCPI-PSI based on the biochemical assay results.


Assuntos
Proteínas de Ligação à Clorofila/metabolismo , Clorofila/metabolismo , Citocromos c6/metabolismo , Diatomáceas/metabolismo , Fragmentos de Peptídeos/metabolismo , Complexo de Proteína do Fotossistema I/metabolismo , Clorofila/química , Clorofila A , Proteínas de Ligação à Clorofila/química , Cromatografia Líquida , Citocromos c6/química , Diatomáceas/citologia , Fluorescência , Fragmentos de Peptídeos/química , Fotoquímica , Complexo de Proteína do Fotossistema I/química , Espectrometria de Massas em Tandem
8.
J Phys Chem B ; 128(3): 731-743, 2024 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-38198639

RESUMO

The exciton states on the smallest type-I photosynthetic reaction center complex of a green sulfur bacterium Chlorobaculum tepidum (GsbRC) consisting of 26 bacteriochlorophylls a (BChl a) and four chlorophylls a (Chl a) located on the homodimer of two PscA reaction center polypeptides were investigated. This analysis involved the study of exciton states through a combination of theoretical modeling and the genetic removal of BChl a pigments at eight sites. (1) A theoretical model of the pigment assembly exciton state on GsbRC was constructed using Poisson TrESP (P-TrESP) and charge density coupling (CDC) methods based on structural information. The model reproduced the experimentally obtained absorption spectrum, circular dichroism spectrum, and excitation transfer dynamics, as well as explained the effects of mutation. (2) Eight BChl a molecules at different locations on the GsbRC were selectively removed by genetic exchange of the His residue, which ligates the central Mg atom of BChl a, with the Leu residue on either one or two PscAs in the RC. His locations are conserved among all type-I RC plant polypeptide, cyanobacteria, and bacteria amino acid sequences. (3) Purified mutant-GsbRCs demonstrated distinct absorption and fluorescence spectra at 77 K, which were different from each other, suggesting successful pigment removal. (4) The same mutations were applied to the constructed theoretical model to analyze the outcomes of these mutations. (5) The combination of theoretical predictions and experimental mutations based on structural information is a new tool for studying the function and evolution of photosynthetic reaction centers.


Assuntos
Chlorobi , Cianobactérias , Complexo de Proteínas do Centro de Reação Fotossintética , Complexo de Proteínas do Centro de Reação Fotossintética/química , Chlorobi/química , Mutação , Cianobactérias/metabolismo , Enxofre/metabolismo , Bacterioclorofilas/química , Proteínas de Bactérias/química
9.
Biochemistry ; 52(26): 4451-9, 2013 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-23750703

RESUMO

Drought-tolerant mosses survive with their green color intact even after long periods of dehydration that would kill ordinary plants. The mechanism of dissipation of excitation energy under drought stress was studied in two species of drought-tolerant moss, Rhytidium rugosum and Ceratodon purpureus. They showed severe quenching of photosystem II chlorophyll fluorescence (PSII) after being dehydrated in the dark. Quenching was induced by the acceleration of the fluorescence decay rate. This drought-induced nonphotochemical quenching (designated d-NPQ) was fully reversed by rehydration. Global analysis of fluorescence decay at 77 K indicated rapid 46 ps transfer of excitation energy from the 680-690 nm PSII bands to a 710 nm band, and to 740-760 nm bands. The latter bands decayed to the ground state with the same time constant showing the rapid dissipation of excitation energy into heat. The quenching by d-NPQ in dry moss was stronger than that by PSII charge separation or nonphotochemical quenching (NPQ), which operates under hydrating conditions. Drought-tolerant mosses, thus, dissipate excess excitation energy into heat. The d-NPQ mechanism in moss resembles that reported in lichens, suggesting their common origin.


Assuntos
Bryopsida/química , Clorofila/química , Metabolismo Energético/fisiologia , Complexo de Proteína do Fotossistema II/metabolismo , Bryopsida/fisiologia , Clorofila/metabolismo , Clorofila/fisiologia , Dessecação , Secas , Fluorescência , Luz , Complexos de Proteínas Captadores de Luz , Complexo de Proteínas do Centro de Reação Fotossintética , Complexo de Proteína do Fotossistema II/genética
10.
Biochim Biophys Acta ; 1817(2): 287-97, 2012 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-22037395

RESUMO

Functional roles of an anionic lipid phosphatidylglycerol (PG) were studied in pgsA-gene-inactivated and cdsA-gene-inactivated/phycobilisome-less mutant cells of a cyanobacterium Synechocystis sp. PCC 6803, which can grow only in PG-supplemented media. 1) A few days of PG depletion suppressed oxygen evolution of mutant cells supported by p-benzoquinone (BQ). The suppression was recovered slowly in a week after PG re-addition. Measurements of fluorescence yield indicated the enhanced sensitivity of Q(B) to the inactivation by BQ. It is assumed that the loss of low-affinity PG (PG(L)) enhances the affinity for BQ that inactivates Q(B). 2) Oxygen evolution without BQ, supported by the endogenous electron acceptors, was slowly suppressed due to the direct inactivation of Q(B) during 10 days of PG depletion, and was recovered rapidly within 10h upon the PG re-addition. It is concluded that the loss of high-affinity PG (PG(H)) displaces Q(B) directly. 3) Electron microscopy images of PG-depleted cells showed the specific suppression of division of mutant cells, which had developed thylakoid membranes attaching phycobilisomes (PBS). 4) Although the PG-depletion for 14 days decreased the chlorophyll/PBS ratio to about 1/4, flourescence spectra/lifetimes were not modified indicating the flexible energy transfer from PBS to different numbers of PSII. Longer PG-depletion enhanced allophycocyanin fluorescence at 683nm with a long 1.2ns lifetime indicating the suppression of energy transfer from PBS to PSII. 5) Action sites of PG(H), PG(L) and other PG molecules on PSII structure are discussed.


Assuntos
Fosfatidilgliceróis/metabolismo , Complexo de Proteína do Fotossistema II/química , Complexo de Proteína do Fotossistema II/metabolismo , Plastoquinona/química , Plastoquinona/metabolismo , Benzoquinonas/antagonistas & inibidores , Benzoquinonas/metabolismo , Sítios de Ligação/efeitos dos fármacos , Catálise/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Cristalografia por Raios X , Transporte de Elétrons/efeitos dos fármacos , Modelos Biológicos , Modelos Moleculares , Organismos Geneticamente Modificados , Oxigênio/metabolismo , Oxigênio/farmacologia , Fosfatidilgliceróis/química , Fosfatidilgliceróis/farmacologia , Pigmentos Biológicos/química , Pigmentos Biológicos/metabolismo , Ligação Proteica/efeitos dos fármacos , Synechocystis/citologia , Synechocystis/efeitos dos fármacos , Synechocystis/genética , Synechocystis/metabolismo
11.
Plant Cell Physiol ; 54(8): 1316-25, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23737501

RESUMO

Lichens are drought-resistant symbiotic organisms of mycobiont fungi and photobiont green algae or cyanobacteria, and have an efficient mechanism to dissipate excess captured light energy into heat in a picosecond time range to avoid photoinhibition. This mechanism can be assessed as drought-induced non-photochemical quenching (d-NPQ) using time-resolved fluorescence spectroscopy. A green alga Trebouxia sp., which lives within a lichen Ramalina yasudae, is one of the most common green algal photobionts. This alga showed very efficient d-NPQ under desiccation within the lichen thallus, whereas it lost d-NPQ ability when isolated from R. yasudae, indicating the importance of the interaction with the mycobiont for d-NPQ ability. We analyzed the water extracts from lichen thalli that enhanced d-NPQ in Trebouxia. Of several sugar compounds identified in the water extracts by nuclear magnetic resonance (NMR), mass spectrometry (MS) and gas chromatography (GC) analyses, only d-arabitol recovered d-NPQ in isolated Trebouxia to a level similar to that detected for R. yasudae thallus. Other sugar compounds did not help the expression of d-NPQ at the same concentrations. Thus, arabitol is essential for the expression of d-NPQ to dissipate excess captured light energy into heat, protecting the photobiont from photoinhibition. The relationship between mycobionts and photobionts is, therefore, not commensalism, but mutualism with each other, as shown by d-NPQ expression.


Assuntos
Ascomicetos/fisiologia , Clorófitas/fisiologia , Líquens/fisiologia , Álcoois Açúcares/metabolismo , Simbiose , Clorofila/metabolismo , Clorófitas/efeitos da radiação , Dessecação , Fluorescência , Líquens/microbiologia , Líquens/efeitos da radiação , Luz
12.
Langmuir ; 29(17): 5104-9, 2013 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-23590586

RESUMO

LH1-α and -ß polypeptides, which make up the light-harvesting 1 (LH1) complex of purple photosynthetic bacteria, along with bacteriochlorophylls, have unique binding properties even for various porphyrin analogs. Herein, we used the porphyrin analogs, Zn-Chlorin and the Zn-Chlorin dimer, and examined their binding behaviors to the LH1-α variant, which has a His-tag at the C-terminus (MBP-rubα-YH). Zn-Chlorin and the Zn-Chlorin dimer could bind to MBP-rubα-YH and form a subunit-type assembly, similar to that from the native LH1 complex. These complexes could be immobilized onto Ni-nitrilotriacetic acid-modified Au electrodes, and the cathodic photocurrent was successfully observed by photoirradiation. Since Zn-Chlorins in this complex are too far for direct electron transfer from the electrode, a contribution of polypeptide backbone for efficient electron transfer was implied. These findings not only show interesting properties of LH1-α polypeptides but also suggest a clue to construct artificial photosynthesis systems using these peptide materials.


Assuntos
Clorofila/biossíntese , Clorofila/química , Ouro/química , Histidina/metabolismo , Proteínas Imobilizadas/metabolismo , Complexos de Proteínas Captadores de Luz/metabolismo , Zinco/química , Eletrodos , Histidina/química , Proteínas Imobilizadas/biossíntese , Proteínas Imobilizadas/química , Complexos de Proteínas Captadores de Luz/química , Estrutura Molecular , Peptídeos/química , Peptídeos/metabolismo , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo
13.
J Bacteriol ; 194(24): 6828-36, 2012 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-23043001

RESUMO

The moderately halotolerant cyanobacterium Synechocystis sp. strain PCC 6803 contains a plasma membrane aquaporin, AqpZ. We previously reported that AqpZ plays a role in glucose metabolism under photomixotrophic growth conditions, suggesting involvement of AqpZ in cytosolic osmolarity homeostasis. To further elucidate the physiological role of AqpZ, we have studied its gene expression profile and its function in Synechocystis. The expression level of aqpZ was regulated by the circadian clock. AqpZ activity was insensitive to mercury in Xenopus oocytes and in Synechocystis, indicating that the AqpZ can be categorized as a mercury-insensitive aquaporin. Stopped-flow light-scattering spectrophotometry showed that addition of sorbitol and NaCl led to a slower decrease in cell volume of the Synechocystis ΔaqpZ strain than the wild type. The ΔaqpZ cells were more tolerant to hyperosmotic shock by sorbitol than the wild type. Consistent with this, recovery of oxygen evolution after a hyperosmotic shock by sorbitol was faster in the ΔaqpZ strain than in the wild type. In contrast, NaCl stress had only a small effect on oxygen evolution. The amount of AqpZ protein remained unchanged by the addition of sorbitol but decreased after addition of NaCl. This decrease is likely to be a mechanism to alleviate the effects of high salinity on the cells. Our results indicate that Synechocystis AqpZ functions as a water transport system that responds to daily oscillations of intracellular osmolarity.


Assuntos
Aquaporinas/metabolismo , Pressão Osmótica , Synechocystis/fisiologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Transporte Biológico/genética , Tamanho Celular , Relógios Circadianos , Perfilação da Expressão Gênica , Regulação Bacteriana da Expressão Gênica , Cloreto de Mercúrio/farmacologia , Concentração Osmolar , Oxigênio/metabolismo , Cloreto de Sódio/farmacologia , Sorbitol/farmacologia , Synechocystis/genética , Água/metabolismo
14.
Biochim Biophys Acta ; 1807(7): 803-12, 2011 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-21420930

RESUMO

The 6xHis-tag-pscA gene, which was genetically engineered to express N-terminally histidine (His)-tagged PscA, was inserted into a coding region of the recA gene in the green sulfur bacterium Chlorobaculum tepidum (C. tepidum). Although the inactivation of the recA gene strongly suppressed a homologous recombination in C. tepidum genomic DNA, the mutant grew well under normal photosynthetic conditions. The His-tagged reaction center (RC) complex could be obtained simply by Ni(2+)-affinity chromatography after detergent solubilization of chlorosome-containing membranes. The complex consisted of three subunits, PscA, PscB, and PscC, in addition to the Fenna-Matthews-Olson protein, but there was no PscD. Low-temperature EPR spectroscopic studies in combination with transient absorption measurements indicated that the complex contained all intrinsic electron transfer cofactors as detected in the wild-type strain. Furthermore, the LC/MS/MS analysis revealed that the core protein consisted of a mixture of a His-/His-tagged PscA homodimer and a non-/His-tagged PscA heterodimer. The development of the pscA gene duplication method presented here, thus, enables not only a quick and large-scale preparation of the RC complex from C. tepidum but also site-directed mutagenesis experiments on the artificially incorporated 6xHis-tag-pscA gene itself, since the expression of the authentic PscA/PscA homodimeric RC complex could complement any defect in mutated His-tagged PscA. This method would provide an invaluable tool for structural and functional analyses of the homodimeric type 1 RC complex.


Assuntos
Proteínas de Bactérias/química , Chlorobi/química , Chlorobi/metabolismo , Complexo de Proteínas do Centro de Reação Fotossintética/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Chlorobi/genética , Espectroscopia de Ressonância de Spin Eletrônica , Engenharia Genética , Espectrometria de Massas , Mutagênese Sítio-Dirigida/métodos , Fenótipo , Complexo de Proteínas do Centro de Reação Fotossintética/genética , Complexo de Proteínas do Centro de Reação Fotossintética/metabolismo , Complexo de Proteína do Fotossistema I/química , Complexo de Proteína do Fotossistema I/genética , Complexo de Proteína do Fotossistema I/metabolismo , Conformação Proteica , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Multimerização Proteica , Subunidades Proteicas/química , Subunidades Proteicas/genética , Subunidades Proteicas/metabolismo , Recombinases Rec A/genética , Recombinases Rec A/metabolismo , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo
15.
Plant Cell Physiol ; 53(8): 1492-506, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-22739509

RESUMO

The fluorescence spectrum at 298 and 40 K and the absorption spectrum at 298 K of each cell of the filamentous cyanobacterium Nostoc sp. was measured by single-cell confocal laser spectroscopy to study the differentiation of cell pigments. The fluorescence spectra of vegetative (veg) and heterocyst (het) cells of Nostoc formed separate groups with low and high PSII to PSI ratios, respectively. The fluorescence spectra of het cells at 40 K still contained typical PSII bands. The PSII/PSI ratio estimated for the veg cells varied between 0.4 and 1.2, while that of het cells varied between 0 and 0.22 even in the same culture. The PSII/PSI ratios of veg cells resembled each other more closely in the same filament. 'pro-het' cells, which started to differentiate into het cells, were identified from the small but specific difference in the PSII/PSI ratio. The allophycocyanin (APC)/PSII ratio was almost constant in both veg and het cells, indicating their tight couplings. Phycocyanin (PC) showed higher fluorescence in most het cells, suggesting the uncoupling from PSII. Veg cells seem to vary their PSI contents to give different PSII/PSI ratios even in the same culture, and to suppress the synthesis of PSII, APC and PC to differentiate into het cells. APC and PC are gradually liberated from membranes in het cells with the uncoupling from PSII. Single-cell spectrometry will be useful to study the differentiation of intrinsic pigments of cells and chloroplasts, and to select microbes from natural environments.


Assuntos
Nostoc/citologia , Nostoc/metabolismo , Pigmentos Biológicos/análise , Análise de Célula Única/métodos , Espectrometria de Fluorescência/métodos , Temperatura Baixa , Interpretação Estatística de Dados , Congelamento , Nostoc/crescimento & desenvolvimento , Complexo de Proteína do Fotossistema I/metabolismo , Complexo de Proteína do Fotossistema II/metabolismo , Ficocianina/metabolismo
16.
J Exp Bot ; 63(10): 3765-75, 2012 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-22438303

RESUMO

Three different types of non-photochemical de-excitation of absorbed light energy protect photosystem II of the sun- and desiccation-tolerant moss Rhytidium rugosum against photo-oxidation. The first mechanism, which is light-induced in hydrated thalli, is sensitive to inhibition by dithiothreitol. It is controlled by the protonation of a thylakoid protein. Other mechanisms are activated by desiccation. One of them permits exciton migration towards a far-red band in the antenna pigments where fast thermal deactivation takes place. This mechanism appears to be similar to a mechanism detected before in desiccated lichens. A third mechanism is based on the reversible photo-accumulation of a radical that acts as a quencher of excitation energy in reaction centres of photosystem II. On the basis of absorption changes around 800 nm, the quencher is suggested to be an oxidized chlorophyll. The data show that desiccated moss is better protected against photo-oxidative damage than hydrated moss. Slow drying of moss thalli in the light increases photo-protection more than slow drying in darkness.


Assuntos
Briófitas/enzimologia , Briófitas/efeitos da radiação , Metabolismo Energético/efeitos da radiação , Complexo de Proteína do Fotossistema II/metabolismo , Água/metabolismo , Briófitas/genética , Briófitas/fisiologia , Clorofila/metabolismo , Dessecação , Luz , Oxirredução/efeitos da radiação , Complexo de Proteína do Fotossistema II/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo
18.
Biotechnol Appl Biochem ; 59(6): 451-6, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-23586954

RESUMO

Underwater high-voltage discharges (3.5 kV) resulting in 4.9 kJ shock waves (50-60 MPa) were studied at the laboratory scale as a Jatropha curcas L. seed disintegration method. Grinding and macerating in an excess of methanol (3.5:1) was advantageous because methanol acts both as a liquid carrier for the pressure shock waves and as a solvent that increases the efficiency of oil extraction while remaining usable for esterification. The influence of the number of shock waves and the intensity of methanol maceration on the heat values of the pressed cake are stated in detail. Soxhlet extraction demonstrated that a greater than 94% oil extraction was achieved. The increased disintegration of vacuoles rich in oil was documented by surface area analysis, mineralization kinetics analysis, and electron microscopy. The working volumes were small, and the proportion of energy inadequate compared to the yields released; however, much can be improved by upgrading the process.


Assuntos
Biocombustíveis , Eletricidade , Jatropha/metabolismo , Água
19.
J Obstet Gynaecol Res ; 38(3): 483-9, 2012 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-22381101

RESUMO

AIM: A low-protein diet (LPD) during pregnancy induces vascular dysfunction and hypertension in the offspring, prevented by administration of an angiotensin II type 1 (AT(1)) receptor antagonist in early life to the offspring. Whether such protection extends to subsequent pregnancy is unknown; we therefore hypothesized that administration of a specific AT(1) receptor antagonist (losartan) in early life to offspring of LPD dams would improve vascular dysfunction in their uterine arteries when they, in turn, were pregnant. METHODS: Pregnant rats were randomly divided into two dietary groups fed a control (C) or protein-restricted (R) diet throughout pregnancy. Between two and 10 weeks postnatally, female offspring (F(1)) were randomly assigned to drink either pure tap water (CO, RO) or water with losartan (CL, RL). Offspring were mated and killed on gestational day 19 or 20 in order to investigate uterine artery function. RESULTS: In pregnant offspring, vasoconstriction of the uterine arteries to phenylephrine (PE) and the thromboxane A2 mimetic U46619 was greater in RO than CO (F(1)). Responses to both antagonists were suppressed in RL (F(1)). Relaxation to sodium nitroprusside was increased in RO versus CO and suppressed in RL versus RO (F(1)). CONCLUSION: Administration of an AT(1) receptor antagonist to offspring during the suckling and juvenile period improves the uterine vascular dysfunction in pregnancy induced by prior maternal LPD during their development. Such treatment may contribute to decreasing the transmitted risks of maternal malnutrition from offspring to the subsequent generation.


Assuntos
Bloqueadores do Receptor Tipo 1 de Angiotensina II/uso terapêutico , Dieta com Restrição de Proteínas/efeitos adversos , Losartan/uso terapêutico , Doenças Vasculares Periféricas/prevenção & controle , Complicações Cardiovasculares na Gravidez/prevenção & controle , Artéria Uterina/efeitos dos fármacos , Ácido 15-Hidroxi-11 alfa,9 alfa-(epoximetano)prosta-5,13-dienoico/farmacologia , Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Animais , Feminino , Hipertensão/etiologia , Hipertensão/prevenção & controle , Losartan/farmacologia , Nitroprussiato/farmacologia , Doenças Vasculares Periféricas/etiologia , Doenças Vasculares Periféricas/fisiopatologia , Fenilefrina/farmacologia , Gravidez , Complicações Cardiovasculares na Gravidez/etiologia , Complicações Cardiovasculares na Gravidez/fisiopatologia , Distribuição Aleatória , Ratos , Ratos Wistar , Artéria Uterina/fisiopatologia , Vasoconstrição/efeitos dos fármacos , Vasoconstritores/farmacologia , Vasodilatação/efeitos dos fármacos , Vasodilatadores/farmacologia
20.
Juntendo Iji Zasshi ; 68(2): 140-146, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-38912278

RESUMO

Objective: the primary objective was to examine the effect of Bifidobacterium on decreasing the bilirubin level in term neonates delivered by Caesarean Section (CS). Materials and Methods: A total of 153 healthy term neonates delivered by CS were included in this study and were divided into the non-probiotic group (n=99) and probiotic group (n=54) based on the history of probiotics administration. There were no infants who underwent phototherapy. A total of 20 doses of probiotics were given orally from the first day of life. The transcutaneous bilirubin (TcB) levels were measured every day for the first 5 days of life. Data of each infant and mother were gathered from medical records. Results: The bilirubin level per day (day-1 to day-5) in the non-probiotic group was no different from the probiotic group. Differences in bilirubin level between day-5 and day-1, and also between day-5 and day-2 were not different between the two groups. There was a significant (p = 0.03) body weight gain in the probiotic groups with a mean of 36.09 ± 8.23 gram/day. No obvious adverse reactions were seen in both the non-probiotic group and probiotic group. Conclusions: Our findings suggest no significant effects of probiotics on lowering bilirubin levels in the first five days of life. Also, probiotics have a positive effect on body weight gain in healthy term infants, and it is safe to be given to newborns.

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