Characterization, feasibility and optimization of Agaricus subrufescens growth based on chemical elements on casing layer

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Partial purification and characterization of lysine-ketoglutarate reductase in normal and opaque-2 maize endosperms

Lysine-ketoglutaratc reductase catalyzes the first step of lysine catabolism in maize (Zea mays L.) endosperm. The enzyme condenses L-lysine and α-ketoglutarate into saccharopine using NADPH as cofactor. It is endosperm-specific and has a temporal pattern of activity, increasing with the onset of kernel development, reaching a peak 20 to 25 days after pollination, and thereafter decreasing as the kernel approaches maturity. The enzyme was extracted from the developing maize endosperm and partially purified by ammonium-sulfate precipitation, anion-exchange chromatography on DEAE-cellulose, and affinity chromatography on Blue-Sepharose CL-6B. The preparation obtained from affinity chromatography was enriched 275-fold and had a specific activity of 411 nanomoles per minute per milligram protein. The native and denaturated enzyme is a 140 kilodalton protein as determined by polyacrylamide gel electrophoresis. The enzyme showed specificity for its substrates and was not inhibited by either aminoethyl-cysteine or glutamate. Steady-state product-inhibition studies revealed that saccharopine was a noncompetitive inhibitor with respect to α-ketoglutarate and a competitive inhibitor with respect to lysine. This is suggestive of a rapid equilibriumordered binding mechanism with a binding order of lysine, α-ketoglutarate, NADPH. The enzyme activity was investigated in two maize inbred lines with homozygous normal and opaque-2 endosperms. The pattern of lysine-ketoglutarate reductase activity is coordinated with the rate of zein accumulation during endosperm development. A coordinated regulation of enzyme activity and zein accumulation was observed in the opaque-2 endosperm as the activity and zein levels were two to three times lower than in the normal endosperm. Enzyme extracted from L1038 normal and opaque-2 20 days after pollination was partially purified by DEAE-cellulose chromatography. Both genotypes showed a similar elution pattern with a single activity peak eluted at approximately 0.2 molar KCL. The molecular weight and physical properties of the normal and opaque-2 enzymes were essentially the same. We suggest that the Opaque-2 gene, which is a transactivator of the 22 kilodalton zein genes, may be involved in the regulation of the lysine-ketoglutarate reductase gene in maize endosperm. In addition, the decreased reductase activity caused by the opaque-2 mutation may explain, at least in part, the elevated concentration of lysine found in the opaque-2 endosperm.

Níveis críticos de fósforo no solo e nas folhas para a implantação de Eucalyptus grandis Hill ex Maiden, em quatro tipos de solos

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

Purification and characterization of two β-glucosidases from the thermophilic fungus Thermoascus aurantiacus

β-Glucosidase from the fungus Thermoascus aurantiacus grown on semi-solid fermentation medium (using ground corncob as substrate) was partially purified in 5 steps-ultrafiltration, ethanol precipitation, gel filtration and 2 anion exchange chromatography runs, and characterized. After the first anion exchange chromatography, β-glucosidase activity was eluted in 3 peaks (Gl-1, Gl-2, Gl-3). Only the Gl-2 and Gl-3 fractions were adsorbed on the gel matrix. Gl-2 and Gl-3 exhibited optimum pH at 4.5 and 4.0, respectively. The temperature optimum of both glucosidases was at 75-80°C. The pH stability of Gl-2 (4.0-9.0) was higher than Gl-3 (5.5-8.5); both enzyme activities showed similar patterns of thermostability. Under conditions of denaturing gel chromatography the molar mass of Gl-2 and Gl-3 was 175 and 157 kDa, respectively. Using 4-nitrophenyl β-D-glucopyranoside as substrate, Km values of 1.17 ± 0.35 and 1.38 ± 0.86 mmol/L were determined for Gl-2 and Gl-3, respectively. Both enzymes were inhibited by Ag+ and stimulated by Ca2+.

One-step purification of 5-enolpyruvylshikimate-3-phosphate synthase enzyme from Mycobacterium tuberculosis

Currently, there are 8 million new cases and 2 million deaths annually from tuberculosis, and it is expected that a total of 225 million new cases and 79 million deaths will occur between 1998 and 2030. The reemergence of tuberculosis as a public health threat, the high susceptibility of HIV-infected persons, and the proliferation of multi-drug-resistant strains have created a need to develop new antimycobacterial agents. The existence of homologues to the shikimate pathway enzymes has been predicted by the determination of the genome sequence of Mycobacterium tuberculosis. We have previously reported the cloning and overexpression of M. tuberculosis aro A-encoded EPSP synthase in both soluble and active forms, without IPTG induction. Here, we describe the purification of M. tuberculosis EPSP synthase (mtEPSPS) expressed in Escherichia coli BL21(DE3) host cells. Purification of mtEPSPS was achieved by a one-step purification protocol using an anion exchange column. The activity of the homogeneous enzyme was measured by a coupled assay using purified shikimate kinase and purine nucleoside phosphorylase proteins. A total of 53 mg of homogeneous enzyme could be obtained from 1 L of LB cell culture, with a specific activity value of approximately 18 U mg-1. The results presented here provide protein in quantities necessary for structural and kinetic studies, which are currently underway in our laboratory. © 2002 Elsevier Science (USA). All rights reserved.

Efeitos do fósforo do solo no desenvolvimento inicial de Genipa americana L.

A greenhouse experiment was carried out to evaluate the effects of soil phosphorus in the development of plants of Genipa americana L. Five doses of P (0, 50, 100, 200, and 400 mg dm -3) were applied in pots with 10 dm 3 of three Neossolos Quartzarênicos (NQ 1, NQ 2 and NQ 3) and one Latossolo Vermelho (LV), that were used in an experiment with Eucalyptus plants during three months. After this period, the soils were analyzed and a 30 cm height seedling of G. americana was planted in each pot. The experiment design was completely randomized, in 5 × 4 factorial scheme, with five replications. The largest biomass of the stem was obtained with 45 mg dm -3 of P in the soil NQ 1 (9% of clay) and 59 mg dm -3 in the soil NQ 3 (14% of clay). In the LV (40% of clay), the biomass of the stem increased linearly in the band of 3 to 148 mg dm -3 of P. The phosphorus critical level in the soil for 80% of the steam and leaves dry matter production of G. americana at planting phase is 15 mg dm -3, using an anion-exchange resin as extractor.

cDNA cloning and 1.75 Å crystal structure determination of PPL2, an endochitinase and N-acetylglucosamine-binding hemagglutinin from Parkia platycephala seeds

Parkia platycephala lectin 2 was purified from Parkia platycephala (Leguminosae, Mimosoideae) seeds by affinity chromatography and RP-HPLC. Equilibrium sedimentation and MS showed that Parkia platycephala lectin 2 is a nonglycosylated monomeric protein of molecular mass 29 407 ± 15 Da, which contains six cysteine residues engaged in the formation of three intramolecular disulfide bonds. Parkia platycephala lectin 2 agglutinated rabbit erythrocytes, and this activity was specifically inhibited by N-acetylglucosamine. In addition, Parkia platycephala lectin 2 hydrolyzed β(1-4) glycosidic bonds linking 2-acetoamido-2-deoxy-β-d-glucopyranose units in chitin. The full-length amino acid sequence of Parkia platycephala lectin 2, determined by N-terminal sequencing and cDNA cloning, and its three-dimensional structure, established by X-ray crystallography at 1.75 Å resolution, showed that Parkia platycephala lectin 2 is homologous to endochitinases of the glycosyl hydrolase family 18, which share the (βα) 8 barrel topology harboring the catalytic residues Asp125, Glu127, and Tyr182. © 2006 The Authors.

Propriedades químicas de um latossolo vermelho de cerrado cultivado com algodão sob diferentes preparos e adubações

The high rate of mineralization of organic matter on savannah soils, which is reached fi ve times faster than in temperate regions, leads us to the challenge of electing the best system of management that maintains and/or increase it in soil, guaranteeing its quality and sustainability. In this sense the present research aimed to study the effects of green, organic and mineral manure on the chemical properties on the chemical properties of an Oxisol, on Savannah area, cultivated with cotton (Gossypium hirsutum) under conventional tillage and no-tillage in the pasture fi eld (Brachiaria decumbens) for 20 years. The experimental design was a randomized block design with split plots. The main plots consisted of two treatments: conventional tillage and no-tillage, and the subplots of six treatments: control (no fertilizer), mineral fertilizer recommended for the crop, according to the soil chemical analysis, organic fertilizer (cattle manure - 20 t ha-1), organic fertilizer (cattle manure - 20 t ha-1) + 1/2mineral fertilization recommended according to the analysis of soil, green manure-1 (Crotalaria juncea) and green manure-2 (Pennisetum americanum). There were studied the following soil chemical properties: P, OM, pH, K, Ca, Mg, Al, Al + H, S, exchange capacity cations and base saturation. The soil samples for the analysis were performed on layers of 0,00-0,05 m, 0,05-0,10 and 0,10-0,20 m. Then it came the following conclusions: the fertilization interfere in soil chemical properties and the preparation did not interfere, the cattle manure and its association with the mineral fertilizer caused increasing level elements in the soil, Crotalaria juncea and Pennisetum americanum did not infl uence on soil chemical properties.

Atributos químicos do solo sob plantio direto afetado por sistemas de culturas e fontes de adubação

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Indicators of soil degradation in urban forests: Physical and chemical parameters

The fragmentation of forest habitats in urban areas has aroused increasing interest in recent years according to the growing environmental problems. The fragmentation of theses ecosystems is caused, in general, by the pressure of housing, agriculture and industry, causing losses in biodiversity and problems of soil degradation in the border areas of theses remnants. The establishment of indicators of soil degradation becomes essential for the implementation of conservation and reclamation. This study analyzes physical and chemical characteristics of soil under different forms of vegetation in the forest surrounding the Quilombo Forest, located in Campinas/SP - Brazil, and examines the possibility of using these indices as indicators of environmental degradation in urban remnants. The parameters analyzed were: specific weight natural (γn), specific weight of solids (γs) Ca, P, K, Mg, pH, organic matter, H + Al, Sum of Base (SB) Percent Base Saturation (V%), Cation Exchange Capacity (CEC). The study shows that in general the different forms of land used in the study area significantly changed (or according to) the physical aspects of soil The porosity and voids of the soil stood out as the best indicators of soil physical degradation in the layer 0-20 cm deep. In relation to chemical indices, the soil under the cultivation of cane sugar had a significantly higher pH, K, Ca, Mg and sum of bases. The areas of forest showed higher levels of phosphorus, organic matter and CEC, indicating the importance of maintaining vegetation and replacement for the cycling of organic matter.

Recovery of degraded areas using topsoil in the Amazon rainforest

This study is aimed at evaluating the effect of placing the top soil cover areas, such as tailings degraded by tin mining, in the Amazon rainforest in Brazil. The evaluations of the planting sites occurred in areas where tin mining was carried out, basically planting native trees over a period of seven years. This work did not come from a pre-decreed methodology of experimental design, and data was collected only seven years after planting. Thus, it was not possible to identify all variables that contributed to a better recovery of the areas. Sampling was done about seven years after placing the top soil and is determined: pH, organic matter content, P, K, Ca, Mg, Al, cation exchange capacity (T), base saturation (V%), B, Fe, Zn, Cu and Mn. The native forest species existing at the site were evaluated in relation to height (in meters) and diameter of the base. For all sites where the surface layer of soil was applied, there were significant differences in the growth of native species. A fundamental aspect in the rehabilitation of areas degraded by mining, in general, is the knowledge about the soil where that recovery must be conducted. The specific procedures in the rehabilitation of those areas depend essentially on the physical, chemical, biological and mineralogical properties of the soil, which must present conditions for the adequate development of the plants. The initial idea of implanting a project of recovery of soils degraded by mining in the Amazon Forest emerged from a first visit to the field, carried out in 1998. The conditions of the already mined areas, in comparison to the exuberant forest of the surroundings caught our attention. The mining company that acts in the area had already been trying for some time to implant a plan of recovery of these mining areas, however without reaching any significant positive results. The loss of organic matter is one of the main problems of degraded areas in Brazil. The storage and reuse of a blanket of soil (topsoil) produce excellent results, but most of the miners consider this technique expensive and difficult because of operational costs and the sharp topographical condition of the mine site. Therefore, a research project was elaborated prioritizing the recovery of the soil degraded by the tin mining as a prior step to the recovery activities with native forest species. The formation of a superficial pseudo-horizon that supported the vegetation and the time that it would take for its establishment became the main objective of this research. The objective of this work is to verify the levels of elements and their traces in areas where top soils were applied for the remediation of degraded areas with local re-vegetation. © 2011 WIT Press.

Impacts of agricultural activities in remaining forest: Campinas/SP, Brazil

Forest fragmentation occurs normally in an area around the city or with high agricultural influence, such as the Forest of Quilombo that lies in Metropolitan Campinas/SP- Brazil. This forest is one such example since it is separated from the other forest fragments in the region for several types of human action. The objective of this study is to analyze the macro and micronutrients and soil edaphic insect fauna in the forest, pasture and sugar cane and inferring the impacts caused by the Mata do Quilombo cattle farming and urban expansion in chemical aspects of soil. Samples were collected in June/11, according to the procedure of method traps pitfall traps. In addition, at each sampling point four composite samples were collected for soil fertility analysis. Samples were collected at six points: pasture, degraded forest (near pasture), preserved forest (near pasture), degraded forest (near cane sugar), preserved forest (near cane sugar), and sugar cane sugar. The samples thus prepared were analyzed Ca, P, K, Mg, pH, organic matter, H + Al, Sum of Base (SB), Base Percentage Saturation (V%), Cation Exchange Capacity (CEC) and trace elements (S, B, Cu, Fe, Mg and Zn). Generally it can be seen that the group of organisms of soil fauna presented with little biodiversity. The number of individuals also shows little species, taxonomic groups showing the highest degree of impact that the remaining forest has suffered. Regarding the analysis of fertility it can be observed that the soil of the surrounding areas of the forest is under direct influence of agriculture. © 2013 WIT Press.

Influence of Agroforestry and Traditional Management of Maize (Zea mays L.) on Soil Fertility

The aim of the work was to evaluate soil nutrient concentration at 0-5, 5-10, and 10-20 cm in maize (Zea mays L.) grown in sequence with black oats (Avena strigosa Schreb.) under Leucaena diversifolia alley cropping agroforestry system (AFS) and traditional management system/sole crop (without trees-TS), following a randomized block design. The experiment was carried out at the Brazilian Association of Biodynamic Agriculture, in Botucatu, São Paulo, Brazil. The treatments were: control (C), chemical fertilizer (F), biomass of L. diversifolia alley cropping (B), and biomass of L. diversifolia alley cropping + chemical fertilizer (B+F). After 2 yr, it was observed that pH, organic matter, and nutrient content had a tendency to show higher values in the treatments biomass+fertilizer, biomass, and fertilizer application, in both systems. Higher values in pH, organic matter, phosphorus, potassium, calcium, magnesium, sum of bases, cation exchange capacity, percentage base saturation, boron, copper, and manganese tended to occur in the agroforestry system. © 2013 Copyright Taylor and Francis Group, LLC.

Caracterização da fertilidade do solo da composição mineral de espécies arbóreas de restinga do litoral paulista

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Atributos de latossolos e produtividade da cana-de-açúcar em decorrência da diversificação de culturas no período de reforma do canavial

Pós-graduação em Agronomia (Ciência do Solo) - FCAV