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Braz. j. microbiol ; 47(1): 143-149, Jan.-Mar. 2016. tab, graf
Artigo em Inglês | LILACS | ID: lil-775118


Abstract Enzyme production by Aspergillus terreus NCFT 4269.10 was studied under liquid static surface and solid-state fermentation using mustard oil cake as a substrate. The maximum lipase biosynthesis was observed after incubation at 30 °C for 96 h. Among the domestic oils tested, the maximum lipase biosynthesis was achieved using palm oil. The crude lipase was purified 2.56-fold to electrophoretic homogeneity, with a yield of 8.44%, and the protein had a molecular weight of 46.3 kDa as determined by SDS-PAGE. Enzyme characterization confirmed that the purified lipase was most active at pH 6.0, temperature of 50 °C, and substrate concentration of 1.5%. The enzyme was thermostable at 60 °C for 1 h, and the optimum enzyme–substrate reaction time was 30 min. Sodium dodecyl sulfate and commercial detergents did not significantly affect lipase activity during 30-min incubation at 30 °C. Among the metal ions tested, the maximum lipase activity was attained in the presence of Zn2+, followed by Mg2+ and Fe2+. Lipase activity was not significantly affected in the presence of ethylenediaminetetraacetic acid, sodium lauryl sulfate and Triton X-100. Phenylmethylsulfonyl fluoride (1 mM) and the reducing, β-mercaptoethanol significantly inhibited lipase activity. The remarkable stability in the presence of detergents, additives, inhibitors and metal ions makes this lipase unique and a potential candidate for significant biotechnological exploitation.

Aspergillus/enzimologia , Lipase/metabolismo , Cátions Bivalentes/metabolismo , Eletroforese em Gel de Poliacrilamida , Estabilidade Enzimática , Ativadores de Enzimas/análise , Inibidores Enzimáticos/análise , Concentração de Íons de Hidrogênio , Lipase/química , Lipase/isolamento & purificação , Peso Molecular , Mercaptoetanol/metabolismo , Metais/metabolismo , Temperatura
Braz. j. microbiol ; 44(2): 559-567, 2013. tab
Artigo em Inglês | LILACS | ID: lil-688589


One variable at a time procedure was used to evaluate the effect of qualitative variables on the production of tannase from Aspergillus niger Van Tieghem. These variables including: fermentation technique, agitation condition, tannins source, adding carbohydrates incorporation with tannic acid, nitrogen source type and divalent cations. Submerged fermentation under intermittent shaking gave the highest total tannase activity. Maximum extracellular tannase activity (305 units/ 50 mL) was attained in medium containing tannic acid as tannins source and sodium nitrate as nitrogen source at 30 ºC for 96 h. All added carbohydrates showed significant adverse effects on the production of tannase. All tested divalent cations significantly decreased tannase production. Moreover, split plot design was carried out to study the effect of fermentation temperature and fermentation time on tannase production. The results indicated maximum tannase production (312.7 units/50 mL) at 35 ºC for 96 h. In other words, increasing fermentation temperature from 30 ºC to 35 ºC resulted in increasing tannase production.

Aspergillus niger/enzimologia , Hidrolases de Éster Carboxílico/metabolismo , Cátions Bivalentes/metabolismo , Meios de Cultura/química , Inibidores Enzimáticos/metabolismo , Fermentação , Temperatura , Fatores de Tempo
Neotrop. entomol ; 36(1): 65-69, Jan.-Feb. 2007. graf
Artigo em Português | LILACS | ID: lil-447094


As ATPases, um importante alvo de inseticidas, são enzimas que hidrolisam o ATP e utilizam a energia liberada no processo para realizar algum tipo de trabalho celular. A larva de Pachymerus nucleorum (Fabricius) possui uma ATPase que apresenta alta atividade Ca-ATPásica, mas não expressa atividade Mg-ATPásica. Nesse trabalho, foi testado o efeito de íons zinco e cobre na atividade Ca-ATPásica dessa enzima. Mais de 90 por cento da atividade Ca-ATPásica foi inibida em 0,5 mM de íons cobre ou 0,25 mM de íons zinco. Na presença de EDTA, mas não na sua ausência, a inibição por zinco foi revertida pelo aumento da concentração de cálcio. A inibição por íons cobre, não foi revertida nem na presença e nem na ausência de EDTA. O tratamento da fração ATPase com cobre, previamente ao ensaio de atividade ATPásica, não inibiu a atividade Ca-ATPásica sugerindo que o íon cobre não liga diretamente a enzima. Os resultados sugerem que íons zinco e cobre formam complexo com o ATP e se ligam à enzima inibindo sua atividade Ca-ATPásica.

ATPases, an important target of insecticides, are enzymes that hydrolyze ATP and use the energy released in that process to accomplish some type of cellular work. Pachymerus nucleorum (Fabricius) larvae possess an ATPase, that presents high Ca-ATPase activity, but no Mg-ATPase activity. In the present study, the effect of zinc and copper ions in the activity Ca-ATPase of that enzyme was tested. More than 90 percent of the Ca-ATPase activity was inhibited in 0.5 mM of copper ions or 0.25 mM of zinc ions. In the presence of EDTA, but not in the absence, the inhibition by zinc was reverted with the increase of calcium concentration. The inhibition by copper ions was not reverted in the presence or absence of EDTA. The Ca-ATPase was not inhibited by treatment of the ATPase fraction with copper, suggesting that the copper ion does not bind directly to the enzyme. The results suggest that zinc and copper ions form a complex with ATP and bind to the enzyme inhibiting its Ca-ATPase activity.

Animais , Besouros/enzimologia , Besouros/crescimento & desenvolvimento , ATPases Transportadoras de Cálcio/efeitos dos fármacos , ATPases Transportadoras de Cálcio/metabolismo , Cobre/farmacologia , Zinco/farmacologia , Cátions Bivalentes/farmacologia , Larva/enzimologia
Biol. Res ; 39(1): 173-182, 2006. ilus
Artigo em Inglês | LILACS | ID: lil-430710


Based on indirect evidence, a role for synaptically released copper and zinc as modulators of neuronal activity has been proposed. To test this proposal directly, we studied the effect of copper, zinc, and other divalent cations on voltage-dependent currents in dissociated toad olfactory neurons and on their firing rate induced by small depolarizing currents. Divalent cations in the nanomolar range sped up the activation kinetics and increased the amplitude of the inward sodium current. In the micromolar range, they caused a dose dependent inhibition of the inward Na+ and Ca2+ currents (INa and ICa) and reduced de amplitude of the Ca2+-dependent K+ outward current (ICa-K). On the other hand, the firing rate of olfactory neurons increased when exposed to nanomolar concentration of divalent cations and decreased when exposed to micromolar concentrations. This biphasic effect of divalent cations on neuronal excitability may be explained by the interaction of these ions with high and low affinity sites in voltage-gated channels. Our results support the idea that these ions are normal modulators of neuronal excitability.

Animais , Cobre/farmacologia , Neurônios Receptores Olfatórios/efeitos dos fármacos , Zinco/farmacologia , Anuros , Potenciais de Ação/efeitos dos fármacos , Potenciais de Ação/fisiologia , Membrana Celular , Cátions Bivalentes/farmacologia , Estimulação Elétrica , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Neurônios Receptores Olfatórios/fisiologia , Transdução de Sinais/fisiologia
Biol. Res ; 36(3/4): 367-379, 2003. ilus, tab, graf
Artigo em Inglês | LILACS | ID: lil-356880


Trypanosoma cruzi epimastigotes were extracted under various conditions in order to examine the role of divalent cations in the solubilization of microtubule proteins. When epimastigotes were homogenized in the presence of 5 mM Mg+2 and 5 mM Ca+2, a protein kinase responsible for phosphorylating tubulin, as well as the tubulin that became phosphorylated, remained tightly associated with the parasite particulate and detergent-resistant fractions. On the contrary, tubulin kinase and its substrate were predominantly released into the parasite cytosolic and detergent-soluble fractions, when epimastigotes were extracted in the presence of 5 mM EDTA and 5 mM EGTA. These evidences demonstrated a divalent cation-dependent solubilization of the enzyme responsible for the phosphorylation of tubulin in T. cruzi epimastigotes and suggested a tight association between tubulin and this kinase. Under all conditions tested, tubulin kinase activity in epimastigote extracts was lower than the addition of the corresponding value in the parasite cytosolic and membranous fractions, suggesting the presence of a kinase inhibitor or regulatory subunit which also seemed to be modulated by divalent cations. Additionally, inhibition experiments in the presence of heparin, 2,3-bisphosphoglycerate and GTP established that the parasite tubulin kinase corresponded to a protein kinase CK2.

Animais , Proteína Quinase C , Trypanosoma cruzi , Tubulina (Proteína) , Cátions Bivalentes , Eletroforese em Gel de Poliacrilamida , Fosforilação , Solubilidade
Braz. j. med. biol. res ; 25(1): 53-5, 1992. ilus
Artigo em Inglês | LILACS | ID: lil-109000


The marine sponge Anthosigmella varians contains proteins that agglutinate human erythrocytes irrespective of their ABO group antigens. The hemagglutination reaction depends on divalent cations andf is not inhibited by L-arabinose, D-xylose, L-rhamnose, D-galactose, D-glucose, L and D-fucose, N-acetyl-D-galactosamine, N-acetyl-D-glucosamine, methyl-alpha-Dmannopiranoside, D-cellobiose, lactose, maltose, melibiose nor raffinose (33 mM each). A partial purification of the hemagglutinins with 31-fold ioncrease in SA and 80% recovery of activity was obtained after gel filtration and ion-exchange gradient elution chromatography. Hemadsorption experiments carried our with out with the semipurified fraction using glutaraldehyde-fixed human erythrocytes suggest that protein with molecular weight of 90 and 34 kDa participate in this rection

Animais , Hemaglutininas/análise , Poríferos/química , Brasil , Cátions Bivalentes , Cromatografia por Troca Iônica , Hemadsorção , Água do Mar
Braz. j. med. biol. res ; 20(6): 759-61, 1987. tab
Artigo em Inglês | LILACS | ID: lil-77431


The aminopeptidase activity of rats urine was tested using L-aminoacy1-2-naphtylamides and L-Leu-p-nitroanilide. The enzyme preparation was purified by ion-exchange and gel filtration chromatography and showed only a single active protein active protein band on 7.5% polyacrylamide gel electrophoresis. The enzyme was characterized by studying the effects of ions (divalent cations and chloride), chelating agents and -SH and -S-S- group reagents and by determining kinetic parameters (Km and Vmax for different substrates and K1 for amino acids, antibiotics and anti-inflammatory drugs)

Ratos , Animais , Aminopeptidases/antagonistas & inibidores , Aminopeptidases/urina , Aminopeptidases/isolamento & purificação , Aminopeptidases/metabolismo , Cátions Bivalentes/farmacologia , Cinética