Effects of temperature and heat treatment time of ground soybeans on the rate of ammonia release in vitro

O trabalho foi conduzido no Laboratório de Nutrição Animal da Faculdade de Ciências Agrárias e Veterinárias (UNESP), Campus de Jaboticabal, com o objetivo de determinar os efeitos da temperatura (0,60°C, 90°C e 120°C) e do tempo de aquecimento (15, 40 e 80 minutos) de grãos de soja moídos, sobre a taxa de liberação de amônia em ensaio de fermentação in vitro, o grau de solubilidade da proteína e a concentração de nitrogênio não-protéico. Foram utilizados seis tratamentos com três repetições em delineamento inteiramente casualizado. Os teores médios de amônia variaram de 28,82 mg a 100,68 mg de nitrogênio amoniacal/100 ml de fluido ruminal, a solubilidade da proteína de 28,60% a 31,22%, e a concentração de nitrogênio não-protéico de 0,49% a 0,62%.This research was conducted in the Laboratory of the Department of Animal Nutrition at the Faculdade de Ciências Agrárias e Veterinárias (UNESP), "Campus", in Jaboticabal, SP, Brazil. This work aimed at determining the effects of temperature (0.60°C, 90°C and 120°C) and heat treatment time (15,40 and 80 minutes) of ground soybeans on the rate of ammonia release in an in vitro fermentation experiment, the rate of protein solubility and the non-protein nitrogen concentration. Six treatments with three replicates in a totally random statistical design were utilized. Ammonia contents ranged from 28.82 mg to 100.68 mg NH3 - N/100 ml of ruminal liquid, the protein solubility ranged from 28.6% to 31.22%, and the concentration of non-protein nitrogen ranged from 0.49% to 0.62%

Pervasive gaps in Amazonian ecological research

Biodiversity loss is one of the main challenges of our time,1,2 and attempts to address it require a clear un derstanding of how ecological communities respond to environmental change across time and space.3,4 While the increasing availability of global databases on ecological communities has advanced our knowledge of biodiversity sensitivity to environmental changes,5–7 vast areas of the tropics remain understudied.8–11 In the American tropics, Amazonia stands out as the world’s most diverse rainforest and the primary source of Neotropical biodiversity,12 but it remains among the least known forests in America and is often underrepre sented in biodiversity databases.13–15 To worsen this situation, human-induced modifications16,17 may elim inate pieces of the Amazon’s biodiversity puzzle before we can use them to understand how ecological com munities are responding. To increase generalization and applicability of biodiversity knowledge,18,19 it is thus crucial to reduce biases in ecological research, particularly in regions projected to face the most pronounced environmental changes. We integrate ecological community metadata of 7,694 sampling sites for multiple or ganism groups in a machine learning model framework to map the research probability across the Brazilian Amazonia, while identifying the region’s vulnerability to environmental change. 15%–18% of the most ne glected areas in ecological research are expected to experience severe climate or land use changes by 2050. This means that unless we take immediate action, we will not be able to establish their current status, much less monitor how it is changing and what is being lostinfo:eu-repo/semantics/publishedVersio

Pervasive gaps in Amazonian ecological research

Biodiversity loss is one of the main challenges of our time,1,2 and attempts to address it require a clear understanding of how ecological communities respond to environmental change across time and space.3,4 While the increasing availability of global databases on ecological communities has advanced our knowledge of biodiversity sensitivity to environmental changes,5,6,7 vast areas of the tropics remain understudied.8,9,10,11 In the American tropics, Amazonia stands out as the world's most diverse rainforest and the primary source of Neotropical biodiversity,12 but it remains among the least known forests in America and is often underrepresented in biodiversity databases.13,14,15 To worsen this situation, human-induced modifications16,17 may eliminate pieces of the Amazon's biodiversity puzzle before we can use them to understand how ecological communities are responding. To increase generalization and applicability of biodiversity knowledge,18,19 it is thus crucial to reduce biases in ecological research, particularly in regions projected to face the most pronounced environmental changes. We integrate ecological community metadata of 7,694 sampling sites for multiple organism groups in a machine learning model framework to map the research probability across the Brazilian Amazonia, while identifying the region's vulnerability to environmental change. 15%–18% of the most neglected areas in ecological research are expected to experience severe climate or land use changes by 2050. This means that unless we take immediate action, we will not be able to establish their current status, much less monitor how it is changing and what is being lost

Purification, primary structure and potential functions of a novel lectin from Bauhinia forficata seeds

A new lectin. Bfl. was purified from Bauhinia forficata seeds by ammonium sulfate fractionation. DEAE-Sephadex ion exchange chromatography, Sepharose-4B and chitin affinity chromatographies and Superdex 75 size exclusion chromatography. the molecular homogeneity and purity of BfL were assessed by reversed-phase H PLC. BfL appeared as a single band of approximately 27.0 kDa on SDS-PAGE under non-reducing and reducing conditions, and its molecular weight was determined to be 27,850 Da by LC/ESI-MS. Bit is a glycoprotein with a carbohydrate content of 6.24% determined by the phenol-sulfuric acid method. Fetuin, asialofetuin, thyroglobulin and azocasein inhibited the hemagglutinating activity of BfL, whereas saccharides did not. Bfl hemagglutinating activity was stable at 100 degrees C for 30 min, pH-dependent, with the highest activity at pH 6.0, and metal-independent. the primary structure of BfL shows similarity with other lectins from the genus Bauhinia. Deconvolution of the BfL circular dichroism (CD) spectrum indicated the presence of alpha-helix and beta structures. BfL increases coagulation time, but this effect is not related to human plasma kallikrein or human factor Xa inhibition. Bfl also inhibits ADP- and epinephrine-induced platelet aggregation in a dose-dependent manner and is the only currently described lectin from Bauhinia that exhibits anticoagulant and antiplatelet aggregating properties. (c) 2012 Elsevier B.V. All rights reserved.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Universidade Federal de São Paulo, Dept Bioquim, BR-04044020 São Paulo, BrazilUniversidade Federal de São Paulo, Dept Biofis, BR-04044020 São Paulo, BrazilUniv Estadual Oeste Parana, Ctr Engn & Ciencias Exatas, BR-85903000 Toledo, PR, BrazilUniv Fed Pernambuco, Dept Bioquim, BR-50670901 Recife, PE, BrazilInst Clin Neuroimmunol LMU, Munich, GermanyMax Planck Inst Biochem, Dept Prot Analyt, D-82152 Martinsried, GermanyUniversidade Federal de São Paulo, Dept Bioquim, BR-04044020 São Paulo, BrazilUniversidade Federal de São Paulo, Dept Biofis, BR-04044020 São Paulo, BrazilFAPESP: 07/58929-4FAPESP: 09/53766-5Web of Scienc

Structural characterization of coagulant Moringa oleifera Lectin and its effect on hemostatic parameters

Lectins are carbohydrate recognition proteins. cMoL, a coagulant Moringa oleifera Lectin, was isolated from seeds of the plant. Structural studies revealed a heat-stable and pH resistant protein with 101 amino acids, 11.67 theoretical pI and 81% similarity with a M. oleifera flocculent protein. Secondary structure content was estimated as 46% alpha-helix, 12% beta-sheets, 17% beta-turns and 25% unordered structures belonging to the alpha/beta tertiary structure class. cMoL significantly prolonged the time required for blood coagulation, activated partial thromboplastin (aPTF) and prothrombin times (PT), but was not so effective in prolonging aPTT in asialofetuin presence. cMoL acted as an anticoagulant protein on in vitro blood coagulation parameters and at least on aPTT, the lectin interacted through the carbohydrate recognition domain. (C) 2013 Elsevier B.V. All rights reserved.Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundacao de Amparo a Ciencia e Tecnologia do Estado de Pernambuco (FACEPE)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Univ Fed Pernambuco, Dept Bioquim, BR-50670901 Recife, PE, BrazilUniversidade Federal de São Paulo, Dept Bioquim, BR-04044020 São Paulo, BrazilUniv Estadual Oeste Parana, Ctr Engn & Ciencias Exatas, BR-85903000 Toledo, PR, BrazilLMU, Inst Clin Neuroimmunol, Munich, GermanyMax Planck Inst Biochem, Dept Prot Analyt, D-82152 Martinsried, GermanyUniversidade Federal de São Paulo, Dept Bioquim, BR-04044020 São Paulo, BrazilWeb of Scienc

Crystal Structure of Crataeva tapia Bark Protein (CrataBL) and Its Effect in Human Prostate Cancer Cell Lines

A protein isolated from the bark of Crataeva tapia (CrataBL) is both a Kunitz-type plant protease inhibitor and a lectin. We have determined the amino acid sequence and three-dimensional structure of CrataBL, as well as characterized its selected biochemical and biological properties. We found two different isoforms of CrataBL isolated from the original source, differing in positions 31 (Pro/Leu); 92 (Ser/Leu); 93 (Ile/Thr); 95 (Arg/Gly) and 97 (Leu/Ser). CrataBL showed relatively weak inhibitory activity against trypsin (K-iapp = 43 mu M) and was more potent against Factor Xa (K-iapp = 8.6 mu M), but was not active against a number of other proteases. We have confirmed that CrataBL contains two glycosylation sites and forms a dimer at high concentration. the high-resolution crystal structures of two different crystal forms of isoform II verified the beta-trefoil fold of CrataBL and have shown the presence of dimers consisting of two almost identical molecules making extensive contacts (similar to 645 angstrom(2)). the structure differs from those of the most closely related proteins by the lack of the N-terminal beta-hairpin. in experiments aimed at investigating the biological properties of CrataBL, we have shown that addition of 40 mM of the protein for 48 h caused maximum growth inhibition in MTT assay (47% of DU145 cells and 43% of PC3 cells). the apoptosis of DU145 and PC3 cell lines was confirmed by flow cytometry using Annexin V/FITC and propidium iodide staining. Treatment with CrataBL resulted in the release of mitochondrial cytochrome c and in the activation of caspase-3 in DU145 and PC3 cells

Analysis of caspase-3 activation in prostate cancer cell lines.

<p>DU145 (<b>A</b>) and PC3 (<b>B</b>) (1×10⁵ cells) cell lines were seeded in 6-well plates, following the same protocol for apoptosis with annexin V/FITC and PI staining. Cells treated with CrataBL (40 µM), containing RPMI without FBS were incubated for 48 h at 37°C and 5% (v/v) CO₂. The cells were incubated with 10 µL of cleaved caspase 3 Alexa Fluor 488-conjugated antibody for 40 min and analyzed in FACSCalibur flow cytometer. As control, the cells were treated with medium only. The area in black represents the control and in white, cells treated with CrataBL.</p

A comparison of CrataBL with proteins sharing the β-trefoil fold.

<p>(A) Superimposed regions of the N termini of CrataBL (green) and STI (magenta - PDB ID: 1AVW <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0064426#pone.0064426-Oliva2" target="_blank">[44]</a>). (B) A dimer of CrataBL (red) compared to the obligatory dimers of CNL (yellow - PDB ID: 3NBD <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0064426#pone.0064426-Pohleven1" target="_blank">[9]</a>) and GalNAc/Gal-specific agglutinin from <i>Sclerotinia sclerotiorum</i> (green - PDB ID: 2X2S <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0064426#pone.0064426-Gahloth1" target="_blank">[50]</a>).</p