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1.
Microb Genom ; 8(1)2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-35040428

RESUMO

The development of spots or lesions symptomatic of common scab on root and tuber crops is caused by few pathogenic Streptomyces with Streptomyces scabiei 87-22 as the model species. Thaxtomin phytotoxins are the primary virulence determinants, mainly acting by impairing cellulose synthesis, and their production in S. scabiei is in turn boosted by cello-oligosaccharides released from host plants. In this work we aimed to determine which molecules and which biosynthetic gene clusters (BGCs) of the specialized metabolism of S. scabiei 87-22 show a production and/or a transcriptional response to cello-oligosaccharides. Comparative metabolomic analyses revealed that molecules of the virulome of S. scabiei induced by cellobiose and cellotriose include (i) thaxtomin and concanamycin phytotoxins, (ii) desferrioxamines, scabichelin and turgichelin siderophores in order to acquire iron essential for housekeeping functions, (iii) ectoine for protection against osmotic shock once inside the host, and (iv) bottromycin and concanamycin antimicrobials possibly to prevent other microorganisms from colonizing the same niche. Importantly, both cello-oligosaccharides reduced the production of the spore germination inhibitors germicidins thereby giving the 'green light' to escape dormancy and trigger the onset of the pathogenic lifestyle. For most metabolites - either with induced or reduced production - cellotriose was revealed to be a slightly stronger elicitor compared to cellobiose, supporting an earlier hypothesis which suggested the trisaccharide was the real trigger for virulence released from the plant cell wall through the action of thaxtomins. Interestingly, except for thaxtomins, none of these BGCs' expression seems to be under direct control of the cellulose utilization repressor CebR suggesting the existence of a yet unknown mechanism for switching on the virulome. Finally, a transcriptomic analysis revealed nine additional cryptic BGCs that have their expression awakened by cello-oligosaccharides, suggesting that other and yet to be discovered metabolites could be part of the virulome of S. scabiei.


Assuntos
Vias Biossintéticas/efeitos dos fármacos , Celobiose/farmacologia , Celulose/farmacologia , Tubérculos/microbiologia , Streptomyces/crescimento & desenvolvimento , Trioses/farmacologia , Fatores de Virulência/metabolismo , Perfilação da Expressão Gênica , Regulação Bacteriana da Expressão Gênica/efeitos dos fármacos , Macrolídeos/metabolismo , Metabolômica , Família Multigênica/efeitos dos fármacos , Piperazinas/metabolismo , Tubérculos/crescimento & desenvolvimento , RNA-Seq , Streptomyces/efeitos dos fármacos , Streptomyces/metabolismo , Streptomyces/patogenicidade
3.
Int J Biol Macromol ; 173: 136-145, 2021 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-33482202

RESUMO

Herbivores gastrointestinal microbiota is of tremendous interest for mining novel lignocellulosic enzymes for bioprocessing. We previously reported a set of potential carbohydrate-active enzymes from the metatranscriptome of the Hu sheep rumen microbiome. In this study, we isolated and heterologously expressed two novel glucanase genes, Cel5A-h38 and Cel5A-h49, finding that both recombinant enzymes showed the optimum temperatures of 50 °C. Substrate-specificity determination revealed that Cel5A-h38 was exclusively active in the presence of mixed-linked glucans, such as barley ß-glucan and Icelandic moss lichenan, whereas Cel5A-h49 (EC 3.2.1.4) exhibited a wider substrate spectrum. Surprisingly, Cel5A-h38 initially released only cellotriose from lichenan and further converted it into an equivalent amount of glucose and cellobiose, suggesting a dual-function as both endo-ß-1,3-1,4-glucanase (EC 3.2.1.73) and exo-cellobiohydrolase (EC 3.2.1.91). Additionally, we performed enzymatic hydrolysis of sheepgrass (Leymus chinensis) and rice (Orysa sativa) straw using Cel5A-h38, revealing liberation of 1.91 ± 0.30 mmol/mL and 2.03 ± 0.09 mmol/mL reducing sugars, respectively, including high concentrations of glucose and cellobiose. These results provided new insights into glucanase activity and lay a foundation for bioconversion of lignocellulosic biomass.


Assuntos
Proteínas de Bactérias/metabolismo , Celobiose/biossíntese , Celulose 1,4-beta-Celobiosidase/metabolismo , Endo-1,3(4)-beta-Glucanase/metabolismo , Glucose/biossíntese , Sequência de Aminoácidos , Animais , Proteínas de Bactérias/genética , Celulose/metabolismo , Celulose 1,4-beta-Celobiosidase/genética , Clonagem Molecular , Endo-1,3(4)-beta-Glucanase/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Microbioma Gastrointestinal/fisiologia , Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Glucanos/metabolismo , Hidrólise , Cinética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Rúmen/microbiologia , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Ovinos/microbiologia , Especificidade por Substrato , Trioses/metabolismo , beta-Glucanas/metabolismo
5.
Appl Microbiol Biotechnol ; 104(19): 8327-8337, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32803296

RESUMO

Cellodextrins are non-digestible oligosaccharides that have attracted interest from the food industry as potential prebiotics. They are typically produced through the partial hydrolysis of cellulose, resulting in a complex mixture of oligosaccharides with a varying degree of polymerisation (DP). Here, we explore the defined synthesis of cellotriose as product since this oligosaccharide is believed to be the most potent prebiotic in the mixture. To that end, the cellobiose phosphorylase (CBP) from Cellulomonas uda and the cellodextrin phosphorylase (CDP) from Clostridium cellulosi were evaluated as biocatalysts, starting from cellobiose and α-D-glucose 1-phosphate as acceptor and donor substrate, respectively. The CDP enzyme was shown to rapidly elongate the chains towards higher DPs, even after extensive mutagenesis. In contrast, an optimised variant of CBP was found to convert cellobiose to cellotriose with a molar yield of 73%. The share of cellotriose within the final soluble cellodextrin mixture (DP2-5) was 82%, resulting in a cellotriose product with the highest purity reported to date. Interestingly, the reaction could even be initiated from glucose as acceptor substrate, which should further decrease the production costs.Key points• Cellobiose phosphorylase is engineered for the production of cellotriose.• Cellotriose is synthesised with the highest purity and yield to date.• Both cellobiose and glucose can be used as acceptor for cellotriose production.


Assuntos
Cellulomonas , Glucosiltransferases , Celobiose , Celulose , Glucosiltransferases/genética , Glucosiltransferases/metabolismo , Especificidade por Substrato , Trioses
6.
J Biochem ; 168(3): 243-256, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32330257

RESUMO

Biological degradation of cellulose from dead plants in nature and plant biomass from agricultural and food-industry waste is important for sustainable carbon recirculation. This study aimed at searching diverse cellulose-degrading systems of wild filamentous fungi and obtaining fungal lines useful for cellooligosaccharide production from agro-industrial wastes. Fungal lines with cellulolytic activity were screened and isolated from stacked rice straw and soil in subtropical fields. Among 13 isolated lines, in liquid culture with a nutrition-limited cellulose-containing medium, four lines of Aspergillus spp. secreted 50-60 kDa proteins as markedly dominant components and gave clear activity bands of possible endo-ß-1,4-glucanase in zymography. Mass spectroscopy (MS) analysis of the dominant components identified three endo-ß-1,4-glucanases (GH5, GH7 and GH12) and two cellobiohydrolases (GH6 and GH7). Cellulose degradation by the secreted proteins was analysed by LC-MS-based measurement of derivatized reducing sugars. The enzymes from the four Aspergillus spp. produced cellobiose from crystalline cellulose and cellotriose at a low level compared with cellobiose. Moreover, though smaller than that from crystalline cellulose, the enzymes of two representative lines degraded powdered rice straw and produced cellobiose. These fungal lines and enzymes would be effective for production of cellooligosaccharides as cellulose degradation-intermediates with added value other than glucose.


Assuntos
Aspergillus/enzimologia , Secreções Corporais/enzimologia , Celulase/biossíntese , Celulose 1,4-beta-Celobiosidase/biossíntese , Meios de Cultura/química , Proteínas Fúngicas/biossíntese , Nutrientes , Aspergillus/genética , Biodegradação Ambiental , Celobiose/biossíntese , Celulose/biossíntese , Celulose 1,4-beta-Celobiosidase/genética , Hidrólise , Oligossacarídeos/biossíntese , Oryza/microbiologia , Microbiologia do Solo , Trioses/biossíntese
7.
Acta Crystallogr D Struct Biol ; 75(Pt 12): 1138-1147, 2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31793907

RESUMO

The catalytic domain (residues 128-449) of the Orpinomyces sp. Y102 CelC7 enzyme (Orp CelC7) exhibits cellobiohydrolase and cellotriohydrolase activities. Crystal structures of Orp CelC7 and its cellobiose-bound complex have been solved at resolutions of 1.80 and 2.78 Å, respectively. Cellobiose occupies subsites +1 and +2 within the active site of Orp CelC7 and forms hydrogen bonds to two key residues: Asp248 and Asp409. Furthermore, its substrate-binding sites have both tunnel-like and open-cleft conformations, suggesting that the glycoside hydrolase family 6 (GH6) Orp CelC7 enzyme may perform enzymatic hydrolysis in the same way as endoglucanases and cellobiohydrolases. LC-MS/MS analysis revealed cellobiose (major) and cellotriose (minor) to be the respective products of endo and exo activity of the GH6 Orp CelC7.


Assuntos
Proteínas de Bactérias/química , Celobiose/metabolismo , Celulase/química , Celulose 1,4-beta-Celobiosidase/química , Celulose/metabolismo , Neocallimastigales/enzimologia , Trioses/metabolismo , beta-Glucosidase/química , Sítios de Ligação , Cristalografia por Raios X/métodos , Modelos Moleculares , Conformação Proteica , Especificidade por Substrato
9.
Sci Rep ; 9(1): 13630, 2019 09 20.
Artigo em Inglês | MEDLINE | ID: mdl-31541154

RESUMO

Cellulases play important roles in the dietary fibre digestion in pigs, and have multiple industrial applications. The porcine intestinal microbiota display a unique feature in rapid cellulose digestion. Herein, we have expressed a cellulase gene, p4818Cel5_2A, which singly encoded a catalytic domain belonging to glycoside hydrolase family 5 subfamily 2, and was previously identified from a metagenomic expression library constructed from porcine gut microbiome after feeding grower pigs with a cellulose-supplemented diet. The activity of purified p4818Cel5_2A was maximal at pH 6.0 and 50 °C and displayed resistance to trypsin digestion. This enzyme exhibited activities towards a wide variety of plant polysaccharides, including cellulosic substrates of avicel and solka-Floc®, and the hemicelluloses of ß-(1 → 4)/(1 → 3)-glucans, xyloglucan, glucomannan and galactomannan. Viscosity, reducing sugar distribution and hydrolysis product analyses further revealed that this enzyme was a processive endo-ß-(1 → 4)-glucanase capable of hydrolyzing cellulose into cellobiose and cellotriose as the primary end products. These catalytic features of p4818Cel5_2A were further explored in the context of a three-dimensional homology model. Altogether, results of this study report a microbial processive endoglucanase identified from the porcine gut microbiome, and it may be tailored as an efficient biocatalyst candidate for potential industrial applications.


Assuntos
Bactérias/isolamento & purificação , Celulase/metabolismo , Celulose/metabolismo , Polissacarídeos/metabolismo , Animais , Bactérias/enzimologia , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Domínio Catalítico , Celobiose/metabolismo , Celulase/química , Celulase/genética , Microbioma Gastrointestinal , Modelos Moleculares , Conformação Proteica , Especificidade por Substrato , Suínos , Trioses/metabolismo
10.
Plant Cell Environ ; 42(12): 3241-3252, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31378950

RESUMO

The triose phosphate utilization (TPU) rate has been identified as one of the processes that can limit terrestrial plant photosynthesis. However, we lack a robust quantitative assessment of TPU limitation of photosynthesis at the global scale. As a result, TPU, and its potential limitation of photosynthesis, is poorly represented in terrestrial biosphere models (TBMs). In this study, we utilized a global data set of photosynthetic CO2 response curves representing 141 species from tropical rainforests to Arctic tundra. We quantified TPU by fitting the standard biochemical model of C3 photosynthesis to measured photosynthetic CO2 response curves and characterized its instantaneous temperature response. Our results demonstrate that TPU does not limit leaf photosynthesis at the current ambient atmospheric CO2 concentration. Furthermore, our results showed that the light-saturated photosynthetic rates of plants growing in cold environments are not more often limited by TPU than those of plants growing in warmer environments. In addition, our study showed that the instantaneous temperature response of TPU is distinct from temperature response of the maximum rate of Rubisco carboxylation. The new formulations of the temperature response of TPU derived in this study may prove useful in quantifying the biochemical limits to terrestrial plant photosynthesis and improve the representation of plant photosynthesis in TBMs.


Assuntos
Atmosfera/química , Dióxido de Carbono/farmacologia , Luz , Fosfatos/metabolismo , Fotossíntese/efeitos da radiação , Folhas de Planta/efeitos da radiação , Trioses/metabolismo , Temperatura
11.
Mol Biol Rep ; 46(5): 5443-5454, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31359382

RESUMO

PoCel12A, PoCel12B, and PoCel12C are genes that encode glycoside hydrolase family 12 (GH12) enzymes in Penicillium oxalicum. PoCel12A and PoCel12B are typical GH12 enzymes that belong to fungal subfamilies 12-1 and 12-2, respectively. PoCel12C contains a low-complexity region (LCR) domain, which is not found in PoCel12A or PoCel12B and independent of fungal subfamily 12-1 or 12-2. Recombinant enzymes (named rCel12A, rCel12B and rCel12C) demonstrate existing diversity in the substrate specificities. Although most members in GH family 12 are typical endoglucanases and preferentially hydrolyze ß-1,4-glucan (e.g., carboxymethylcellulose), recombinant PoCel12A is a non-typical endo-(1-4)-ß-glucanase; it preferentially hydrolyzes mix-linked ß-glucan (barley ß-glucan, ß-1,3-1,4-glucan) and slightly hydrolyzes ß-1,4-glucan (carboxymethylcellulose). Recombinant PoCel12B possesses a significantly high activity against xyloglucan. A specific activity of rCel12B toward xyloglucan (239 µmol/min/mg) is the second-highest value known. Recombinant PoCel12C shows low activity toward ß-glucan, carboxymethylcellulose, or xyloglucan. All three enzymes can degrade phosphoric acid-swollen cellulose (PASC). However, the hydrolysis products toward PASC by enzymes are different: the main hydrolysis products are cellotriose, cellotetraose, and cellobiose for rCel12A, rCel12B, and rCel12C, correspondingly. A synergistic action toward PASC among rCel12A and rCel12B is observed, thereby suggesting a potential application for preparing enzyme cocktails used in lignocellulose hydrolysis.


Assuntos
Glicosídeo Hidrolases/genética , Glicosídeo Hidrolases/metabolismo , Especificidade por Substrato/genética , Celulase/genética , Celulose/análogos & derivados , Glucanos , Glicosídeo Hidrolases/química , Concentração de Íons de Hidrogênio , Hidrólise , Lignina , Penicillium/genética , Penicillium/metabolismo , Filogenia , Tetroses , Trioses , Xilanos , beta-Glucanas/metabolismo
12.
J Exp Bot ; 70(20): 5773-5785, 2019 10 24.
Artigo em Inglês | MEDLINE | ID: mdl-31269202

RESUMO

This study aimed to understand the physiological basis of rice photosynthetic response to C source-sink imbalances, focusing on the dynamics of the photosynthetic parameter triose phosphate utilization (TPU). Here, rice (Oriza sativa L.) indica cultivar IR64 were grown in controlled environment chambers under current ambient CO2 concentration until heading, and thereafter two CO2 treatments (400 and 800 µmol mol-1) were compared in the presence and absence of a panicle-pruning treatment modifying the C sink. At 2 weeks after heading, photosynthetic parameters derived from CO2 response curves, and non-structural carbohydrate content of flag leaf and internodes were measured three to four times of day. Spikelet number per panicle and flag leaf area on the main culm were recorded. Net C assimilation and TPU decreased progressively after midday in panicle-pruned plants, especially under 800 µmol mol-1 CO2. This TPU reduction was explained by sucrose accumulation in the flag leaf resulting from the sink limitation. Taking together, our findings suggest that TPU is involved in the regulation of photosynthesis in rice under elevated CO2 conditions, and that sink limitation effects should be considered in crop models.


Assuntos
Oryza/metabolismo , Trioses/metabolismo , Dióxido de Carbono/metabolismo , Mudança Climática , Fotossíntese/fisiologia , Sacarose/metabolismo
13.
Acta Crystallogr D Struct Biol ; 75(Pt 6): 605-615, 2019 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-31205022

RESUMO

The discovery of new glycoside hydrolases that can be utilized in the chemoenzymatic synthesis of carbohydrates has emerged as a promising approach for various biotechnological processes. In this study, recombinant Ps_Cel5A from Pseudomonas stutzeri A1501, a novel member of the GH5_5 subfamily, was expressed, purified and crystallized. Preliminary experiments confirmed the ability of Ps_Cel5A to catalyze transglycosylation with cellotriose as a substrate. The crystal structure revealed several structural determinants in and around the positive subsites, providing a molecular basis for a better understanding of the mechanisms that promote and favour synthesis rather than hydrolysis. In the positive subsites, two nonconserved positively charged residues (Arg178 and Lys216) were found to interact with cellobiose. This adaptation has also been reported for transglycosylating ß-mannanases of the GH5_7 subfamily.


Assuntos
Proteínas de Bactérias/química , Celulase/química , Celulose/química , Pseudomonas stutzeri/enzimologia , Trioses/química , Celulose/metabolismo , Cristalização , Cristalografia por Raios X/métodos , Escherichia coli , Glicosilação , Especificidade por Substrato , Trioses/metabolismo
14.
J Exp Bot ; 70(6): 1755-1766, 2019 03 27.
Artigo em Inglês | MEDLINE | ID: mdl-30868155

RESUMO

During photosynthesis, plants fix CO2 from the atmosphere onto ribulose-bisphosphate, producing 3-phosphoglycerate, which is reduced to triose phosphates (TPs). The TPs are then converted into the end products of photosynthesis. When a plant is photosynthesizing very quickly, it may not be possible to commit photosynthate to end products as fast as it is produced, causing a decrease in available phosphate and limiting the rate of photosynthesis to the rate of triose phosphate utilization (TPU). The occurrence of an observable TPU limitation is highly variable based on species and especially growth conditions, with TPU capacity seemingly regulated to be in slight excess of typical photosynthetic rates the plant might experience. The physiological effects of TPU limitation are discussed with an emphasis on interactions between the Calvin-Benson cycle and the light reactions. Methods for detecting TPU-limited data from gas exchange data are detailed and the impact on modeling of some physiological effects are shown. Special consideration is given to common misconceptions about TPU.


Assuntos
Fosfatos/metabolismo , Fotossíntese/fisiologia , Fenômenos Fisiológicos Vegetais , Trioses/metabolismo
15.
Appl Microbiol Biotechnol ; 103(3): 1275-1287, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30547217

RESUMO

Lignocellulose feedstock constitutes the most abundant carbon source in the biosphere; however, its recalcitrance remains a challenge for microbial conversion into biofuel and bioproducts. Bacillus licheniformis is a microbial mesophilic bacterium capable of secreting a large number of glycoside hydrolase (GH) enzymes, including a glycoside hydrolase from GH family 9 (BlCel9). Here, we conducted biochemical and biophysical studies of recombinant BlCel9, and its low-resolution molecular shape was retrieved from small angle X-ray scattering (SAXS) data. BlCel9 is an endoglucanase exhibiting maximum catalytic efficiency at pH 7.0 and 60 °C. Furthermore, it retains 80% of catalytic activity within a broad range of pH values (5.5-8.5) and temperatures (up to 50 °C) for extended periods of time (over 48 h). It exhibits the highest hydrolytic activity against phosphoric acid swollen cellulose (PASC), followed by bacterial cellulose (BC), filter paper (FP), and to a lesser extent carboxymethylcellulose (CMC). The HPAEC-PAD analysis of the hydrolytic products demonstrated that the end product of the enzymatic hydrolysis is primarily cellobiose, and also small amounts of glucose, cellotriose, and cellotetraose are produced. SAXS data analysis revealed that the enzyme adopts a monomeric state in solution and has a molecular mass of 65.8 kDa as estimated from SAXS data. The BlCel9 has an elongated shape composed of an N-terminal family 3 carbohydrate-binding module (CBM3c) and a C-terminal GH9 catalytic domain joined together by 20 amino acid residue long linker peptides. The domains are closely juxtaposed in an extended conformation and form a relatively rigid structure in solution, indicating that the interactions between the CBM3c and GH9 catalytic domains might play a key role in cooperative cellulose biomass recognition and hydrolysis.


Assuntos
Bacillus licheniformis/enzimologia , Bacillus licheniformis/metabolismo , Celulase/metabolismo , Glicosídeo Hidrolases/metabolismo , Lignina/metabolismo , Catálise , Celobiose/biossíntese , Celulose/análogos & derivados , Celulose/biossíntese , Glucose/biossíntese , Concentração de Íons de Hidrogênio , Espalhamento a Baixo Ângulo , Tetroses/biossíntese , Trioses/biossíntese , Difração de Raios X
16.
Biochem J ; 475(16): 2637-2652, 2018 08 30.
Artigo em Inglês | MEDLINE | ID: mdl-30049894

RESUMO

Dihydroxyacetone (DHA) is the smallest ketotriose, and it is utilized by many organisms as an energy source. However, at higher concentrations, DHA becomes toxic towards several organisms including the budding yeast Saccharomyces cerevisiae In the present study, we show that DHA toxicity is due to its spontaneous conversion to methylglyoxal (MG) within yeast cells. A mutant defective in MG-metabolizing enzymes (glo1Δgre2Δgre3Δ) exhibited higher susceptibility to DHA. Intracellular MG levels increased following the treatment of glo1Δgre2Δgre3Δ cells with DHA. We previously reported that MG depolarized the actin cytoskeleton and changed vacuolar morphology. We herein demonstrated the depolarization of actin and morphological changes in vacuoles following a treatment with DHA. Furthermore, we found that both MG and DHA caused the morphological change in nucleus, and inhibited the nuclear division. Our results suggest that the conversion of DHA to MG is a dominant contributor to its cytotoxicity.


Assuntos
Actinas/metabolismo , Divisão do Núcleo Celular/efeitos dos fármacos , Citotoxinas/farmacologia , Glioxal , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Trioses/farmacologia , Actinas/genética , Glioxal/análogos & derivados , Glioxal/metabolismo , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética
17.
Plant Physiol ; 176(3): 2496-2514, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29371249

RESUMO

Piriformospora indica, an endophytic root-colonizing fungus, efficiently promotes plant growth and induces resistance to abiotic stress and biotic diseases. P. indica fungal cell wall extract induces cytoplasmic calcium elevation in host plant roots. Here, we show that cellotriose (CT) is an elicitor-active cell wall moiety released by P. indica into the medium. CT induces a mild defense-like response, including the production of reactive oxygen species, changes in membrane potential, and the expression of genes involved in growth regulation and root development. CT-based cytoplasmic calcium elevation in Arabidopsis (Arabidopsis thaliana) roots does not require the BAK1 coreceptor or the putative Ca2+ channels TPC1, GLR3.3, GLR2.4, and GLR2.5 and operates synergistically with the elicitor chitin. We identified an ethyl methanesulfonate-induced mutant (cytoplasmiccalcium elevation mutant) impaired in the response to CT and various other cellooligomers (n = 2-7), but not to chitooligomers (n = 4-8), in roots. The mutant contains a single nucleotide exchange in the gene encoding a poly(A) ribonuclease (AtPARN; At1g55870) that degrades the poly(A) tails of specific mRNAs. The wild-type PARN cDNA, expressed under the control of a 35S promoter, complements the mutant phenotype. Our identification of cellotriose as a novel chemical mediator casts light on the complex P. indica-plant mutualistic relationship.


Assuntos
Arabidopsis/microbiologia , Basidiomycota/fisiologia , Celulose/metabolismo , Exorribonucleases/metabolismo , Simbiose/fisiologia , Trioses/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Cálcio/metabolismo , Exorribonucleases/genética , Regulação da Expressão Gênica de Plantas , Mutação , Plantas Geneticamente Modificadas , /metabolismo , Plântula/metabolismo , Plântula/microbiologia , Transdução de Sinais
18.
Nat Chem ; 9(4): 310-317, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-28338685

RESUMO

Phosphoenol pyruvate is the highest-energy phosphate found in living organisms and is one of the most versatile molecules in metabolism. Consequently, it is an essential intermediate in a wide variety of biochemical pathways, including carbon fixation, the shikimate pathway, substrate-level phosphorylation, gluconeogenesis and glycolysis. Triose glycolysis (generation of ATP from glyceraldehyde 3-phosphate via phosphoenol pyruvate) is among the most central and highly conserved pathways in metabolism. Here, we demonstrate the efficient and robust synthesis of phosphoenol pyruvate from prebiotic nucleotide precursors, glycolaldehyde and glyceraldehyde. Furthermore, phosphoenol pyruvate is derived within an α-phosphorylation controlled reaction network that gives access to glyceric acid 2-phosphate, glyceric acid 3-phosphate, phosphoserine and pyruvate. Our results demonstrate that the key components of a core metabolic pathway central to energy transduction and amino acid, sugar, nucleotide and lipid biosyntheses can be reconstituted in high yield under mild, prebiotically plausible conditions.


Assuntos
Evolução Química , Glicólise , Origem da Vida , Fosfoenolpiruvato/química , Trioses/química , Estrutura Molecular , Fosforilação
19.
J Sci Food Agric ; 97(3): 743-752, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-27145288

RESUMO

BACKGROUND: The structure of ß-glucan influences its use in cereal-based foods and feed. The objective of this study was to determine the effect of environment (E) and genotype (G) on ß-glucan fine structure and its genetic control in two-row spring barley with normal starch characteristics. RESULTS: A population of 89 recombinant inbred lines, derived from the cross of two-row spring barley genotypes Merit × H93174006 (H92076F1 × TR238), was characterized for concentration and structure of grain ß-glucan in two environments. Results showed that concentrations of ß-glucan, DP3, DP4 and DP3 + DP4 were positively correlated with each other, suggesting no preference for DP3 or DP4 subunit production in high- or low-ß-glucan lines. The concentrations of ß-glucan, DP3, DP4 and DP3:DP4 ratios were significantly influenced by genotype and environment. However, only DP3:DP4 ratio showed a significant effect of G × E interaction. Association mapping of candidate markers in 119 barley genotypes showed that marker CSLF6_4105 was associated with ß-glucan concentration, whereas Bmac504 and Bmac211 were associated with DP3:DP4 ratio. Bmac273e was associated with both ß-glucan concentration and DP3:DP4 ratio. CONCLUSION: The grain ß-glucan concentration and DP3:DP4 ratio are strongly affected by genotype and environment. Single-marker analyses suggested that the genetic control of ß-glucan concentration and DP3:DP4 ratio was linked to separate chromosomal regions on barley genome. © 2016 Society of Chemical Industry.


Assuntos
Carboidratos da Dieta/análise , Interação Gene-Ambiente , Glucosiltransferases/metabolismo , Hordeum/química , Proteínas de Plantas/metabolismo , Sementes/química , beta-Glucanas/análise , Alberta , Altitude , Ração Animal/análise , Animais , Sequência de Carboidratos , Celulose/genética , Celulose/metabolismo , Clima , Cruzamentos Genéticos , Carboidratos da Dieta/metabolismo , Marcadores Genéticos , Glucosiltransferases/genética , Hordeum/genética , Hordeum/crescimento & desenvolvimento , Hordeum/metabolismo , Humanos , Valor Nutritivo , Melhoramento Vegetal , Proteínas de Plantas/genética , Sementes/genética , Sementes/crescimento & desenvolvimento , Sementes/metabolismo , Especificidade por Substrato , Tetroses/metabolismo , Trioses/metabolismo , beta-Glucanas/química , beta-Glucanas/metabolismo
20.
Planta ; 243(3): 687-98, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26620947

RESUMO

MAIN CONCLUSION: The triose phosphate use limitation was studied using long-term and short term changes in capacity. The TPU limitation caused increased proton motive force; long-term TPU limitation additionally reduced other photosynthetic components. Photosynthetic responses to CO2 can be interpreted primarily as being limited by the amount or activity of Rubisco or the capacity for ribulose bisphosphate regeneration, but at high rates of photosynthesis a third response is often seen. Photosynthesis becomes insensitive to CO2 or even declines with increasing CO2, and this behavior has been associated with a limitation of export of carbon from the Calvin-Benson cycle. It is often called the triose phosphate use (TPU) limitation. We studied the long-term consequences of this limitation using plants engineered to have reduced capacity for starch or sucrose synthesis. We studied short-term consequences using temperature as a method for changing the balance of carbon fixation capacity and TPU. A long-term and short-term TPU limitation resulted in an increase in proton motive force (PMF) in the thylakoids. Once a TPU limitation was reached, any further increases in CO2 was met with a further increase in the PMF but no increase or little increase in net assimilation of CO2. A long-term TPU limitation resulted in reduced Rubisco and RuBP regeneration capacity. We hypothesize that TPU, Rubisco activity, and RuBP regeneration are regulated so that TPU is normally in slight excess of what is required, and that this results in more effective regulation than if TPU were in large excess.


Assuntos
Carbono/metabolismo , Fotossíntese , Plantas/metabolismo , Ribulose-Bifosfato Carboxilase/metabolismo , Trioses/metabolismo , Dióxido de Carbono/metabolismo , Fosfatos/metabolismo , Temperatura , Tilacoides/metabolismo , Tempo
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