The Rv1712 locus from Mycobacterium tuberculosis H37Rv codes for a functional CMP kinase that preferentially Phosphorylates dCMP

The Mycobacterium tuberculosis cmk gene, predicted to encode a CMP kinase (CMK), was cloned and expressed, and its product was purified to homogeneity. Steady-state kinetics confirmed that M. tuberculosis CMK is a monomer that preferentially phosphorylates CMP and dCMP by a sequential mechanism. A plausible role for CMK is discussed. Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Evaluation of creatine kinase (CK) and aspartate aminotransferase (AST) activities after laparoscopic or conventional ovariectomy in queens

Creatine kinase (CK) and aspartate aminotransferase (AST) are mainly muscle-specific enzymes, which can be associated with muscle tissue damage. The aim of this study was to assess the activities of CK and AST during the postoperative period, after conventional (G1) and videolaparoscopic ovariectomy (G2), in queens. A further group (G3) was subjected to anaesthesia only. Results demonstrate that there were significant differences between groups. The highest levels of CK were recorded in Gl, however at a confidence level of p < 0.05 there was no significant difference between groups during the first 6 hours after surgery. A significant (p < 0.05) increase of CK values was identified between 0h and 3h in both groups (Gl and G2). Regarding AST activity there was no significant variation between groups, but again there was a significant difference between values at 0h and 3h after surgery. In conclusion, ovariectomy performed by videolap-aroscopy seems to cause less muscle damage when compared to the conventional method. © 2009 by Verlag Hans Huber, Hogrefe AG, Bem.

Efeito da suplementação com selênio sobre a concentração sérica de creatina kinase em bovinos

The study, evaluated the addition of different concentrations of Se in mineral mixture affecting creatine kinase (CK) serum concentrations in cattle. 60 male, Nellore cattle, at about 12 months old, were randomly assigned to groups (15 calves/ group), Gc, G3,6, G5,4 or G6,4 (0, 3.6, 5.4, and 6.4 mg Se/bovine/day). The levels of serum CK in the cattle were not affected by neither the interaction selenium concentration x time nor the concentration of supplementation. However, CK levels increased over the experiment irrespective of dietary selenium concentration. In addition, the frequency of animals with CK levels above normal increased (p<0.10) in group G6,4. The concentrations of selenium studied here do not affect serum CK in cattle, but the daily concentration of 6.4 mg selenium is not recommended because it is possibly toxic effect.

UMP kinase from Mycobacterium tuberculosis: Mode of action and allosteric interactions, and their likely role in pyrimidine metabolism regulation

The pyrH-encoded uridine 5′-monophosphate kinase (UMPK) is involved in both de novo and salvage synthesis of DNA and RNA precursors. Here we describe Mycobacterium tuberculosis UMPK (MtUMPK) cloning and expression in Escherichia coli. N-terminal amino acid sequencing and electrospray ionization mass spectrometry analyses confirmed the identity of homogeneous MtUMPK. MtUMPK catalyzed the phosphorylation of UMP to UDP, using ATP-Mg 2+ as phosphate donor. Size exclusion chromatography showed that the protein is a homotetramer. Kinetic studies revealed that MtUMPK exhibits cooperative kinetics towards ATP and undergoes allosteric regulation. GTP and UTP are, respectively, positive and negative effectors, maintaining the balance of purine versus pyrimidine synthesis. Initial velocity studies and substrate(s) binding measured by isothermal titration calorimetry suggested that catalysis proceeds by a sequential ordered mechanism, in which ATP binds first followed by UMP binding, and release of products is random. As MtUMPK does not resemble its eukaryotic counterparts, specific inhibitors could be designed to be tested as antitubercular agents. © 2010 Elsevier Inc. All rights reserved.

High Content Screening of a Kinase-Focused Library Reveals Compounds Broadly-Active against Dengue Viruses

Dengue virus is a mosquito-borne flavivirus that has a large impact in global health. It is considered as one of the medically important arboviruses, and developing a preventive or therapeutic solution remains a top priority in the medical and scientific community. Drug discovery programs for potential dengue antivirals have increased dramatically over the last decade, largely in part to the introduction of high-throughput assays. In this study, we have developed an image-based dengue high-throughput/high-content assay (HT/HCA) using an innovative computer vision approach to screen a kinase-focused library for anti-dengue compounds. Using this dengue HT/HCA, we identified a group of compounds with a 4-(1-aminoethyl)-N-methylthiazol-2-amine as a common core structure that inhibits dengue viral infection in a human liver-derived cell line (Huh-7.5 cells). Compounds CND1201, CND1203 and CND1243 exhibited strong antiviral activities against all four dengue serotypes. Plaque reduction and time-of-addition assays suggests that these compounds interfere with the late stage of viral infection cycle. These findings demonstrate that our image-based dengue HT/HCA is a reliable tool that can be used to screen various chemical libraries for potential dengue antiviral candidates. © 2013 Cruz et al.

Photoprotective and antioxidant effects of Rhubarb: Inhibitory action on tyrosinase and tyrosine kinase activities and TNF-α, IL-1α and α-MSH production in human melanocytes

Background: Exposure to ultraviolet (UV) radiation causes various forms of acute and chronic skin damage, including immunosuppression, inflammation, premature aging and photodamage. Furthermore, it induces the generation of reactive oxygen species, produces proinflammatory cytokines and melanocyte-stimulating hormone (MSH) and increases tyrosinase activity. The aim of this study was to evaluate the potential photoprotective effects of Rheum rhaponticum L. rhizome extract on human UV-stimulated melanocytes.Methods: The effects of Rheum rhaponticum rhizome extract on tyrosine kinase activity, and on interleukin-1α (IL-1α), tumour necrosis factor α (TNF-α), and α-MSH production in human epidermal melanocytes were evaluated under UV-stimulated and non-stimulated conditions. Antioxidant activity was evaluated by lipid peroxidation and 1,1-dyphenyl-2-picryl-hydrazyl (DPPH) assays, while anti-tyrosinase activity was evaluated by the mushroom tyrosinase method.Results: Rheum rhaponticum L. rhizome extract showed in vitro antioxidant properties against lipid peroxidation, free radical scavenging and anti-tyrosinase activities, and inhibited the production of IL-1α, TNF-α, α-MSH, and tyrosine kinase activity in melanocytes subjected to UV radiation.Conclusions: These results support the inclusion of Rheum rhaponticum L. rhizome extract into cosmetic, sunscreen and skin care products for the prevention or reduction of photodamage. © 2013 Silveira et al; licensee BioMed Central Ltd.

Effects of (-)-cubebin (Piper cubeba) on cytotoxicity, mutagenicity and expression of p38 MAP kinase and GSTa2 in a hepatoma cell line

(-)-Cubebin is a lignan extracted from the seeds of the pepper Piper cubeba, a commonly eaten spice with beneficial properties, including trypanocidal, anti-inflammatory, analgesic, anti-proliferative and leishmanicidal activities. Because of its therapeutic potential, we investigated the effects of (-)-cubebin on the cytotoxicity, cell proliferation kinetics, mutagenicity and expression of p38 MAP kinase and glutathione S-transferase a2 (GSTa2) using real-time RT-PCR in Rattus norvegicus hepatoma cells. We found that 280 μM (-)-cubebin was cytotoxic after 24, 48 and 72. h of exposure, but not mutagenic at 0.28 μM, 2.8 μM and 28 μM after 26. h. Similarly, exposure to 0.28 μM, 2.8 μM and 28 μM (-)-cubebin for 24, 48, 72 and 96. h did not alter the cell proliferation kinetics. Cells exposed to 28 μM (-)-cubebin for 24. h did not exhibit changes in p38 MAP kinase and GSTa2 expression, indicating that cellular changes were not induced by extracellular stimuli and that (-)-cubebin is likely not metabolized via this pathway. Our results suggest that high levels of (-)-cubebin should be consumed with caution due to the cytotoxic effect observed at the highest concentration. However, at lower concentrations, no cytotoxic, mutagenic or proliferative effects were observed, providing further evidence of the safety of consuming (-)-cubebin. © 2013 Elsevier Inc.

The Mode of Action of Recombinant Mycobacterium tuberculosis Shikimate Kinase: Kinetics and Thermodynamics Analyses

Tuberculosis remains as one of the main cause of mortality worldwide due to a single infectious agent, Mycobacterium tuberculosis. The aroK-encoded M. tuberculosis Shikimate Kinase (MtSK), shown to be essential for survival of bacilli, catalyzes the phosphoryl transfer from ATP to the carbon-3 hydroxyl group of shikimate (SKH), yielding shikimate-3-phosphate and ADP. Here we present purification to homogeneity, and oligomeric state determination of recombinant MtSK. Biochemical and biophysical data suggest that the chemical reaction catalyzed by monomeric MtSK follows a rapid-equilibrium random order of substrate binding, and ordered product release. Isothermal titration calorimetry (ITC) for binding of ligands to MtSK provided thermodynamic signatures of non-covalent interactions to each process. A comparison of steady-state kinetics parameters and equilibrium dissociation constant value determined by ITC showed that ATP binding does not increase the affinity of MtSK for SKH. We suggest that MtSK would more appropriately be described as an aroL-encoded type II shikimate kinase. Our manuscript also gives thermodynamic description of SKH binding to MtSK and data for the number of protons exchanged during this bimolecular interaction. The negative value for the change in constant pressure heat capacity (ΔCp) and molecular homology model building suggest a pronounced contribution of desolvation of non-polar groups upon binary complex formation. Thermodynamic parameters were deconvoluted into hydrophobic and vibrational contributions upon MtSK:SKH binary complex formation. Data for the number of protons exchanged during this bimolecular interaction are interpreted in light of a structural model to try to propose the likely amino acid side chains that are the proton donors to bulk solvent following MtSK:SKH complex formation. © 2013 Rosado et al.

Serum activity of creatine kinase and aminotransferase aspartate enzymes of horses submitted to muscle biopsy and incremental jump test

The objective was to evaluate serum activity of the enzymes creatine kinase (CK) and aspartate aminotransferase (AST), which are leakage enzymes responsive to muscle injury, of athletic horses that underwent muscle biopsy and incremental jump test (IJT) involving incremental jumps. The animals were grouped as follows: the first group, horses with history of superior performance (SP); the second, with a history of inferior performance (IP); and lastly, a control group (CG). All groups underwent biopsy of the gluteus medius muscle, while groups SP and IP were also submitted to the incremental jump test (IJT) 24 hours after biopsy. The IJT consisted of three stages with 40 jumps each, where jump height increased progressively, from 40 to 60 and last, 80cm. Blood samples were drawn before biopsy, and 6 and 24 hours after the exercise as well. The levels of CK serum activity increased 6 hours after exercise and decreased 24 hours later in all groups, including CG. AST activity did not increase after biopsy and exercise. There was no increase of both enzyme activities that could be attributed to the exercise, possibly due to exercise short duration and/or low intensity. We conclude that the muscle biopsy was able to show that there was enough stimulus to cause CK enzyme leakage into the plasma, and consequent detection of increased serum activity, while the incremental jump test did not.

Práticas do ecodesign no polo de jóias folheadas de Limeira: um estudo de caso

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Estratégia competitiva e o papel da função produção: uma análise em empresas do polo calçadista de Birigui

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Adição experimental de novos materiais às argilas da região do polo cerâmico de Santa Gertrudes (SP)

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

Concepções e práticas dos sujeitos envolvidos no curso de licenciatura em educação do campo no polo de Castanhal/PA

Este trabalho aborda as concepções e práticas dos sujeitos do curso de Licenciatura Plena em Educação do Campo/LPEC desenvolvido no Instituto Federal de Educação/IFPA no polo de Castanhal/Pa, o qual tem por embasamento político-pedagógico o Programa de Educação do Campo/PROCAMPO. Com base no objeto de estudo foram definidos como objetivos identificar a concepção teórico/metodológico do PROCAMPO na formação de educadores do campo; analisar a dinâmica do Curso de LPEC no polo de Castanhal e identificar as contribuições do curso na prática dos educadores do campo. Para alcançar os objetivos propostos o processo investigativo realizou pesquisa bibliográfica, pesquisa documental e pesquisa de campo. Na pesquisa bibliográfica foram revisados autores para dar sustentação teórica sobre educação como prática humanizadora com destaque para Brandão (2007), Saviani (1994), Mészáros (2008), Freire (2011), Caldart (2004), Fernandes (2004), Arroyo (1999) entre outros. A pesquisa documental abrangeu o Projeto Pedagógico do Curso de Licenciatura Plena em Educação do Campo; Minuta Original do MEC/Secad; Edital nº 02, de 23/04/2008 (referente à chamada pública para seleção de projetos de instituições públicas de ensino superior para o PROCAMPO); Edital de 16/04/2009 e Edital nº 026/2010 (ambos correspondentes ao processo seletivo de LPEC do IFPA) e a listagem de alunos matriculados no PROCAMPO IFPA/Castanhal em 2010, entre outros. A pesquisa de campo, de caráter qualitativa, utilizou o estudo de caso e recorte temporal delimitado para o período de 2008 a 2012, em função da implementação do PROCAMPO ter ocorrido no ano inicial do período citado e a realização de entrevistas semiestruturada. Os resultados indicam que a materialização do curso em Castanhal apesar das dificuldades de natureza pedagógica e financeira vem causando efeitos positivos na atuação dos educadores do campo.