Isolamento de fungos produtores de enzimas pectinolíticas

Soil samples collected in the campus, UNESP, Araraquara, SP, were employed to isolate and characterize fungi strains with potential pectinolytic enzymes. These enzymes have arisen great interest due to its increasing application in the food industry. Two hundred forty six strains were isolated based on the appearance of colony on PDA medium, morphology (septate mycelia, nonseptate conidiophore, black conidia, and clublike spore-bearing head), after 48 h of growth at 30°C. Strains were selected in solid medium containing pectin citrus as sole carbon source and 0.5% rutenium red. The characterization of pectinolytic production was performed in solid culture and batch fermentation medium containing pectin citrus. The enzyme pectinolytic production was evaluated at 30°C, without agitation in 100 mL of medium containing 2% pectin citrus, 0.2% ammonium sulphate, 0.2% magnesium sulphate, and 0.05% potassium phosphate. The maximum pectinolytic activity (15U/mL) was observed in the medium after Aspergillus sp CFCF-0492 growth, while Aspergillus sp CFCF-CC1 showed the higher level of the final biomass. The pectinolytic activity is more preserved when the fungi-spores were maintained in agar-Czapeck medium.

Fontes e doses de nitrogênio na nutrição, produção e qualidade do feijoeiro

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Relação entre o excesso de bases da dieta, sua manipulação mediante adição de cátions e anions e o pH urinário e equilíbrio ácido-básico de cães

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Fontes e doses de adubação nitrogenada na atividade microbiana, colonização micorrízica e fertilidade em solo com Brachiaria brizantha

Pós-graduação em Ciência Animal - FMVA

Caracterização físico-química de pectinases extracelulares purificadas de Aspergillus giganteus

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Fontes, doses e parcelamento do nitrogênio em feijoeiro de inverno no sistema plantio direto

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Purificação e caracterização bioquímica da α-glicosidase termoestável produzida por Thermoascus aurantiacus CBMAI 756

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Plantas de cobertura e doses de nitrogênio em pré-semeadura em algodoeiro

Pós-graduação em Agronomia - FEIS

Produção de ligno-hemi-celulases por fermentação em estado sólido e avaliação dos efeitos da aplicação das enzimas na composição química e estrutura do bagaço e da palha de cana

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Manejo da adubação com sulfato de amônio no desempenho produtivo do milho cultivado após aveia e trigo

The time, dose and applied nutrients in corn have a direct effect on its productivity. Therefore, the objective was to study the application of N and S in corn as ammonium sulfate, in succession to wheat and oats and evaluate different forms of fertilizer management. The experiment was conducted in a randomized block design in Oxisol (Hapludox). The five treatments with N, at a dose of 120 kg ha(-1) were applied in 20 plots (5x4), according to the management of fertilizer: T1-N (120 kg ha(-1)) full at sowing, T2-N (120 kg ha(-1)) total coverage; T3-N (40 kg ha(-1)) at sowing and N (80 kg ha(-1)) in coverage; T4-N advance in wheat sowing and sowing oats (120 kg ha(-1)), T5-(control). The S doses were corresponding to their concentrations in the fertilizer. Only wheat received a dose of 24 kg N ha(-1) at sowing all plots and oats received 24 kg N ha(-1) at sowing only the portions related to treatment with anticipation of corn N (T4). We evaluated the biomass production of winter crops (oats and wheat), according to the fertilization at sowing, and also the influence of winter crops and management of ammonium sulfate, the corn yield. The oats produced more dry matter in relation to wheat, positively influencing the corn yield, regardless of fertilizer management. The anticipation of ammonium sulfate, the sowing of oats, was favorable to corn yield, equating to other forms of management of fertilizer. Rotation corn and oats, forms management, ammonium sulphate, at seeding, topdressing or applied in split were equally efficient in corn yields.

Fibrolytic enzyme production of Myceliophthora thermophila M.7.7. using inexpensive carbon sources and mineral nutrients

This study investigated the effect of inexpensive carbon and nitrogen sources on enzyme production by Myceliophthora thermophila M.7.7 in solid-state fermentation. Three kinds of lignocellulosic waste (corn straw, sugarcane bagasse and sugarcane straw) and six nitrogen sources (urea, calcium nitrate, analytical ammonium sulphate, yeast extract, agricultural fertilizer NPK 20-05-20 and fertilizing grade ammonium sulphate) were tested. Some physical-chermical parameters of the fermentation, such as temperature, initial pH and moisture content of the substrate on enzyme production, were evoluated. The maximum activities of xylanase (446.9 U/ml) endoglucanase (94.7 U/ml) and beta-glucosidase (2.8 U/ml) were observed in a mixture of corn straw and wheat bran (1:1 w/w) as the carbon source using fertilizer grade ammonium sulphate as the nitrogen source. This production occurred for an incubation period of 96 h, at 40°C, with initial moisture content of 70% and pH 5.0. These results have significant interest since they could be used for the future production of enzymes in a low-cost industrial process.

Untersuchungen zur rekombinanten Expression und Faltung pflanzlicher Tyrosinasen

In ihrer dualen Funktion als Monophenolhydroxylase (EC 1.14.18.1) und Diphenoloxidase (EC 1.10.3.1) ist die Tyrosinase das Schlüsselenzym der Melanogenese, der Synthese des Melanins, und übernimmt damit quer durch alle Organismenreiche Aufgaben von der Pigmentierung bis hin zu einer Beteiligung an der Immunantwort. Sie zählt, zusammen mit den Catecholoxidasen und Hämocyaninen, zu den Typ-3-Kupfer-Proteinen, die sich durch ein Aktives Zentrum auszeichnen, das in der Lage ist, Sauerstoff und phenolische Substrate reversibel zwischen zwei Kupfer-Ionen zu binden. Bisher konnte weder die Funktion der pflanzlichen Tyrosinase genau identifiziert, noch die Struktur eines solchen Enzyms aufgeklärt werden. Mit dem späteren Ziel, durch eine röntgenkristallographische Analyse die zugrunde liegende strukturelle Ursache der zusätzlichen Monophenolhydroxylase-Aktivität von Tyrosinasen gegenüber reinen Catecholoxidasen ermitteln zu können, wurde in dieser Arbeit ein bakterielles Expressionssystem entwickelt, das zur Herstellung einer rekombinanten Tyrosinase oder Polyphenoloxidase (PPO) aus Spinacia oleracea (Spinat) für die Kristallisation verwendet werden kann. Das rekombinante Protein wurde in Form von Inclusion Bodies isoliert, anhand einer Affinitätschromatographie aufgereinigt und in anschließende Rückfaltungsexperimente eingesetzt. In einer parallelen Versuchsreihe konnte Spinat, aufgrund seiner hohen Tyrosinaseaktivität, als geeignetes Objekt für die Isolation des nativen Enzyms identifiziert werden. Im Anschluss an eine Thylakoidpräparation, Solubilisierung der Thylakoidmembranen und Fällung des Proteins mit Ammoniumsulfat, wurden Experimente zur weiteren Anreicherung der Tyrosinase-Aktivität über eine Anionenaustausch-Chromatographie und zur Etablierung einiger nachfolgender Aufreinigungsschritte durchgeführt.

Negative second virial coefficients as predictors of protein crystal growth: Evidence from sedimentation equilibrium studies that refutes the designation of those light scattering parameters as osmotic virial coefficients

The effects of ammonium sulphate concentration on the osmotic second virial coefficient (B-AA/M-A) for equine serum albumin (pH 5.6, 20 degrees C) have been examined by sedimentation equilibrium. After an initial steep decrease with increasing ammonium sulphate concentration, B-AA/M-A assumes an essentially concentration-independent magnitude of 8-9 ml/g. Such behaviour conforms with the statistical-mechanical prediction that a sufficient increase in ionic strength should effectively eliminate the contributions of charge interactions to B-AA/M-A but have no effect on the covolume contribution (8.4 ml/g for serum albumin). A similar situation is shown to apply to published sedimentation equilibrium data for lysozyme (pH 4.5). Although termed osmotic second virial coefficients and designated as such (B-22), the negative values obtained in published light scattering studies of both systems have been described incorrectly because of the concomitant inclusion of the protein-salt contribution to thermodynamic nonideality of the protein. Those negative values are still valid predictors of conditions conducive to crystal growth inasmuch as they do reflect situations in which there is net attraction between protein molecules. However, the source of attraction responsible for the negative virial coefficient stems from the protein-salt rather than the protein-protein contribution, which is necessarily positive. (c) 2005 Elsevier B.V. All rights reserved.

Surface properties of enterococcus faecalis in relation to infective endocarditis

The effect of growth conditions on both the appearance and the antigenic profile of cells of Enterococcus faecalis was investigated using electron micrographs of ruthenium red stained and sectioned cells and SDS-PAGE and blotting techniques respectively. Three specific antigens of molecular weights 73, 40 and 37 kdaltons were of particular interest being expressed most strongly after growth in serum. This medium was deemed to most closely mimic jn vjvo growth conditions reflecting an environment similar to that which the microorganisms would encounter during bacteraemia, preceding the colonisation of the endocardium and the development of infective endocarditis. The 40 and 37 kdalton antigens were shown by immunoqold labelling to be exposed on the surface of the cells although they did not appear to be connected with the fimbriae shown to exist on some of the E. faecalis cells examined by negative staining. The 73, 40 and 37 kdalton antigens were crudely purified using sarkosyl and ammonium sulphate precipitation, and used as the basis of a serodiagnostic test for E. faecalis endocarditis using an ELISA system. This was tested in a blind trial and the success rates were 94% for positives, 90% for negatives with endocarditis caused by other organisms and 80% for E. faecalis infections other than endocarditis. The binding of E.faecalis cells to the serum proteins fibronectin and albumin was investigated using 125I labelled proteins, followed by Scatchard analysis. This showed that· E.faecalis cells do loosely bind large amounts of both of these proteins, thus surely affecting the way in which the host's immune system perceives the cells. The E.faecalis receptor for fibronectin was partially characterised and appeared to involve protein and/or carbohydrate containing components. but did not involve LTA or the 40 and 37 kdalton species specific antigens.

Enterotoxigenic Escherichia coli infection of pigs: expression, extraction, purification and the adhesive properties of the K88 fimbrial adhesin

The ability of Escherichia coli to express the K88 fimbrial adhesin was satisfactorily indicated by the combined techniques of ELISA, haemagglutination and latex agglutination. Detection of expression by electron microscopy and the ability to metabolize raffinose were unsuitable. Quantitative expression of the K88 adhesin was determined by ELISA. Expression was found to vary according to the E.coli strain examined, media type and form. In general it was found that the total amount was greater, while the amount/cfu was less on agar than in broth cultures. Expression of the K88 adhesin during unshaken batch culture was related to the growth rate and was maximal during late logarithmic to early stationary phase. A combination of heat extraction, ammonium sulphate and isoelectric precipitation was found suitable for both large and small scale preparation of purified K88ab adhesin. Extraction of the K88 adhesin was sensitive to pH and it was postulated that this may affect the site of colonisation of by ETEC in vivo. Results of haemagglutination experiments were consistent with the hypothesis that the K88 receptor present on erythrocytes is composed of two elements, one responsible for the binding of K88ab and K88ac and a second responsible for the binding of the K88ad adhesin. Comparison of the haemagglutinating properties of cell-free and cell-bound K88 adhesin revealed some differences probably indicating a minor conformational change in the K88 adhesin on its isolation. The K88ab adhesin was found to bind to erythrocytes over a wide pH range (PH 4-9) and was inhibited by αK88ab and αK88b antisera. Inhibition of haemagglutination was noted with crude heparin, mannan and porcine gastric mucin, chondrosine and several hexosamines, glucosamine in particular. The most potent inhibitor of haemagglutination was n-dodecyl-β-D-glucopyranoside, one of a series of glucosides found to have inhibitory properties. Correlation between hydrophobicity of glucosides tested and degree of inhibition observed suggested hydrophobic forces were important in the interaction of the K88 adhesin with its receptor. The results of Scatchard and Hill plots indicated that binding of the K88ab adhesin to porcine enterocytes in the majority of cases is a two-step, three component system. The first K88 receptor (or site) had a K2. of 1.59x1014M-1 and a minimum of 4.3x104 sites/enterocyte. The second receptor (or site) had a K2 of 4.2x1012M-1 with a calculated 1.75x105 sites/enterocyte. Attempts to inhibit binding of cell-free K88 adhesin to porcine enterocytes by lectins were unsuccessful. However, several carbohydrates including trehalose, lactulose, galactose 1→4 mannopyranoside, chondrosine, galactosamine, stachyose and mannan were inhibitory. The most potent inhibitor was found to be porcine gastric mucin. Inhibition observed with n-octyl-α-D-glucopyranose was difficult to interpret in isolation because of interference with the assay, however, it agreed with the results of haemagglutination inhibition experiments.