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Jose Roberto Meyerfernandes - One of the best experts on this subject based on the ideXlab platform.
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trypanosoma rangeli differential expression of cell surface polypeptides and ecto phosphatase activity in short and long epimastigote forms
Experimental Parasitology, 2006Co-Authors: S A O Gomes, Andre Luiz Fonseca De Souza, Bianca A Silva, Tina Kiffermoreira, Jacenir Reis Dos Santosmallet, Andre L S Santos, Jose Roberto MeyerfernandesAbstract:Abstract Trypanosoma rangeli is a parasite of a numerous wild and domestic animals, presenting wide geographical distribution and high immunological cross-reactivity with Trypanosoma cruzi, which may lead to misdiagnosis. T. rangeli has a complex life cycle, involving distinct morphological and functional forms in the vector. Here, we characterized the cell surface polypeptides and the phosphatase activities in short and long epimastigotes forms of T. rangeli, using intact living parasites. The surface protein profile revealed by the incubation of parasites with biotin showed a preferential expression of the 97, 70, 50, 45, 25–22, and 15 kDa biotinylated polypeptides in the long forms, in contrast to the 55 and 28 kDa biotinylated polypeptides synthesized by the short epimastigotes. Additionally, flow cytometry analysis showed that the short forms had relatively lower biotin surface binding than long ones. The involvement of phosphatases with the trypanosomatid differentiation has been proposed. In this sense, T. rangeli living parasites were able to hydrolyze the artificial substrate p-nitrophenylphosphate at a rate of 25.57 ± 2.03 and 10.09 ± 0.93 nmol p-NPP × h−1 × 107 cells for the short and long epimastigotes, respectively. These phosphatase activities were linear with time for at least 60 min and the optimum pH lies in the acid range. Classical inhibitors of acid phosphatases, such as ammonium Molybdate, Sodium fluoride, and zinc chloride, showed a significant decrease in these phosphatase activities, with different patterns of inhibition. Additionally, these phosphatase activities presented different kinetic parameters (Km and Vmax) and distinct sensitivities to divalent cations. Both epimastigotes were unable to release phosphatase to the extracellular environment. Cytochemical analysis demonstrated the localization of these enzymes on the parasite surfaces (cell body and flagellum) and in intracellular vacuoles, resembling acidocalcisomes.
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trypanosoma rangeli characterization of a mg dependent ecto atp diphosphohydrolase activity
Experimental Parasitology, 2006Co-Authors: Fabio V Fonseca, Andre Luiz Fonseca De Souza, Ana C Mariano, Peter F Entringer, Katia C Gondim, Jose Roberto MeyerfernandesAbstract:Abstract In this work we describe the ability of living Trypanosoma rangeli to hydrolyze extracellular ATP. In these intact parasites whose viability was assessed before and after the reactions by motility and by Trypan blue dye exclusion, there was a low level of ATP hydrolysis in the absence of any divalent metal (1.53 ± 0.12 nmol P i /h × 10 7 cells). The ATP hydrolysis was stimulated by MgCl 2 and the Mg-dependent ecto-ATPase activity was 5.24 ± 0.64 nmol P i /h × 10 7 cells. The Mg-dependent ecto-ATPase activity was linear with cell density and with time for at least 60 min. This stimulatory effect on the ATP hydrolysis was also observed when MgCl 2 was replaced by MnCl 2 , but not by CaCl 2 , SrCl 2 , and ZnCl 2 . The apparent K m for Mg-ATP2- was 0.53 ± 0.11 mM. The optimum pH for the T. rangeli Mg-dependent ecto-ATPase activity lies in the alkaline range. This ecto-ATPase activity was insensitive to inhibitors of other ATPase and phosphatase activities, such as oligomycin, Sodium azide, bafilomycin A1, ouabain, furosemide, vanadate, Molybdate, Sodium fluoride, tartrate, and levamizole. To confirm that this Mg-dependent ATPase was an ecto-ATPase, we used an impermeant inhibitor, DIDS (4, 4′-diisothiocyanostylbene 2′-2′-disulfonic acid) as well as suramin, an antagonist of P2 purinoreceptors and inhibitor of some ecto-ATPases. These two reagents inhibited the Mg 2+ -dependent ATPase activity in a dose-dependent manner. This ecto-ATPase activity was stimulated by carbohydrates involved in the attachment/invasion of salivary glands of Rhodnius prolixus and by lipophorin, an insect lipoprotein circulating in the hemolymph.
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characterization of an ecto atpase of tritrichomonas foetus
Veterinary Parasitology, 2002Co-Authors: J B Jesus, Angela H C S Lopes, Jose Roberto MeyerfernandesAbstract:Abstract In this work, we describe the ability of living Tritrichomonas foetus to hydrolyze extracellular ATP. The addition of MgCl2 to the assay medium increased the ecto-ATPase activity in a dose-dependent manner. At 5 mM ATP, half maximal stimulation of ATP hydrolysis was obtained with 0.46 mM MgCl2. The ecto-ATPase activity was also stimulated by MnCl2 and CaCl2, but not by SrCl2. The Mg2+-dependent ATPase presents two apparent Km values for Mg-ATP2− (Km1=0.03 mM and Km2=2.01 mM). ATP was the best substrate for this enzyme, although other nucleotides such as ITP, CTP, UTP also produced high reaction rates. GTP produced a low reaction rate and ADP was not a substrate for this enzyme. The Mg2+-dependent ecto-ATPase activity was insensitive to inhibitors of other ATPase and phosphatase activities, such as oligomycin, Sodium azide, bafilomycin A1, ouabain, furosemide, vanadate, Molybdate, Sodium fluoride and levamizole. The acid phosphatase inhibitors (vanadate and Molybdate) inhibited about 60–70% of the Mg2+-independent ecto-ATPase activity, suggesting that the ATP hydrolysis measured in the absence of any metal divalent could, at least in part, also be catalyzed by an ecto-phosphatase present in this cell. In order to confirm the observed Mg2+-dependent activity as an ecto-ATPase, we used an impermeant inhibitor, 4,4′-diisothiocyanostylbene-2′,2′-disulfonic acid (DIDS) as well as suramin, an antagonist of P2 purinoreceptors and inhibitor of some ecto-ATPases. These two reagents inhibited the Mg2+-dependent ATPase activity in a dose-dependent manner. This ecto-ATPase was stimulated by more than 90% by 50 mM d -galactose. Since previous results showed that d -galactose exposed on the surface of host cells is involved with T. foetus adhesion, the Mg2+-dependent ecto-ATPase may be involved with cellular adhesion and possible pathogenicity.
José Meyer-fernandes - One of the best experts on this subject based on the ideXlab platform.
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Ectonucleotide diphosphohydrolase activity in Crithidia deanei
Parasitology Research, 2002Co-Authors: Adriana Dos Passos Lemos, Ana Acacia De Sá Pinheiro, Márcia De Berrêdo-pinho, André Fonseca De Souza, Maria M. Motta, Wanderley De Souza, José Meyer-fernandesAbstract:In this work we describe the ability of living Crithidia deanei to hydrolyze extracellular ATP. In intact cells at pH 7.2, a low level of ATP hydrolysis was observed in the absence of any divalent metal (0.41±0.13 nmol P_i h^–1 10^7 cells^–1). The ATP hydrolysis was stimulated by MgCl_2 and the Mg^2+-dependent ecto-ATPase activity was 4.05±0.17 nmol P_i h^–1 10^7 cells^–1. Mg^2+-dependent ecto-ATPase activity increased linearly with cell density and with time for at least 60 min. The addition of MgCl_2 to extracellular medium increased the ecto-ATPase activity in a dose-dependent manner. At 5 mM ATP, half-maximal stimulation of ATP hydrolysis was obtained with 0.93±0.26 mM MgCl_2. This stimulatory activity was also observed when MgCl_2 was replaced by MnCl_2, but not CaCl_2 or SrCl_2. The apparent K _m for Mg-ATP^2– was 0.26±0.03 mM. ATP was the best substrate for this enzyme; other nucleotides, such as ITP, GTP, UTP and CTP, produced lower reaction rates. In the pH range from 6.6 to 8.4, in which the cells were viable, the acid phosphatase activity also present in this cell decreased, while the Mg^2+-dependent ATPase activity did not change. This ecto-ATPase activity was insensitive to inhibitors of other ATPase and phosphatase activities, such as oligomycin, Sodium azide, bafilomycin A_1, ouabain, vanadate, Molybdate, Sodium fluoride and tartrate. To confirm that this Mg^2+-dependent ATPase was an ecto-ATPase, we used the impermeant inhibitor 4, 4′-diisothiocyanostylbene 2′-2′-disulfonic acid as well as suramin, an antagonist of P_2 purinoreceptors and inhibitor of some ecto-ATPases. These two reagents inhibited the Mg^2+-dependent ATPase activity in a dose-dependent manner. The cell surface location of the ATP-hydrolyzing site was also confirmed by cytochemical analysis.
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A Mg-Dependent Ecto-ATPase in Leishmania amazonensis and Its Possible Role in Adenosine Acquisition and Virulence
Archives of Biochemistry and Biophysics, 2001Co-Authors: Márcia De Berrêdo-pinho, Carlos E. Peres-sampaio, P.p.m. Chrispim, Rodrigo Belmont-firpo, Adriana Lemos, Martiny A, Marcos A. Vannier-santos, José Meyer-fernandesAbstract:Abstract The plasma membrane of cells contains enzymes whose active sites face the external medium rather than the cytoplasm. The activities of these enzymes, referred to as ectoenzymes, can be measured using living cells. In this work we describe the ability of living promastigotes of Leishmania amazonensis to hydrolyze extracellular ATP. In these intact parasites whose viability was assessed before and after the reactions by motility and by trypan blue dye exclusion, there was a low level of ATP hydrolysis in the absence of any divalent metal (5.39 ± 0.71 nmol P i /h × 10 7 cells). The ATP hydrolysis was stimulated by MgCl 2 and the Mg-dependent ecto-ATPase activity was 30.75 ± 2.64 nmol P i /h × 10 7 cells. The Mg-dependent ecto-ATPase activity was linear with cell density and with time for at least 60 min. The addition of MgCl 2 to extracellular medium increased the ecto-ATPase activity in a dose-dependent manner. At 5 mM ATP, half-maximal stimulation of ATP hydrolysis was obtained with 1.21 mM MgCl 2 . This stimulatory activity was also observed when MgCl 2 was replaced by MnCl 2 , but not by CaCl 2 or SrCl 2 . The apparent K m for Mg-ATP 2− was 0.98 mM and free Mg 2+ did not increase the ecto-ATPase activity. In the pH range from 6.8 to 8.4, in which the cells were viable, the acid phosphatase activity decreased, while the Mg 2+ -dependent ATPase activity increased. This ecto-ATPase activity was insensitive to inhibitors of other ATPase and phosphatase activities, such as oligomycin, Sodium azide, bafilomycin A 1 , ouabain, furosemide, vanadate, Molybdate, Sodium fluoride, tartrate, and levamizole. To confirm that this Mg-dependent ATPase was an ecto-ATPase, we used an impermeant inhibitor, 4,4′-diisothiocyanostylbene 2′,2′-disulfonic acid as well as suramin, an antagonist of P 2 purinoreceptors and inhibitor of some ecto-ATPases. These two reagents inhibited the Mg 2+ -dependent ATPase activity in a dose-dependent manner. A comparison between the Mg 2+ -dependent ATPase activity of virulent and avirulent promastigotes showed that avirulent promastigotes were less efficient than the virulent promastigotes in hydrolyzing ATP.
J B Jesus - One of the best experts on this subject based on the ideXlab platform.
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characterization of an ecto atpase of tritrichomonas foetus
Veterinary Parasitology, 2002Co-Authors: J B Jesus, Angela H C S Lopes, Jose Roberto MeyerfernandesAbstract:Abstract In this work, we describe the ability of living Tritrichomonas foetus to hydrolyze extracellular ATP. The addition of MgCl2 to the assay medium increased the ecto-ATPase activity in a dose-dependent manner. At 5 mM ATP, half maximal stimulation of ATP hydrolysis was obtained with 0.46 mM MgCl2. The ecto-ATPase activity was also stimulated by MnCl2 and CaCl2, but not by SrCl2. The Mg2+-dependent ATPase presents two apparent Km values for Mg-ATP2− (Km1=0.03 mM and Km2=2.01 mM). ATP was the best substrate for this enzyme, although other nucleotides such as ITP, CTP, UTP also produced high reaction rates. GTP produced a low reaction rate and ADP was not a substrate for this enzyme. The Mg2+-dependent ecto-ATPase activity was insensitive to inhibitors of other ATPase and phosphatase activities, such as oligomycin, Sodium azide, bafilomycin A1, ouabain, furosemide, vanadate, Molybdate, Sodium fluoride and levamizole. The acid phosphatase inhibitors (vanadate and Molybdate) inhibited about 60–70% of the Mg2+-independent ecto-ATPase activity, suggesting that the ATP hydrolysis measured in the absence of any metal divalent could, at least in part, also be catalyzed by an ecto-phosphatase present in this cell. In order to confirm the observed Mg2+-dependent activity as an ecto-ATPase, we used an impermeant inhibitor, 4,4′-diisothiocyanostylbene-2′,2′-disulfonic acid (DIDS) as well as suramin, an antagonist of P2 purinoreceptors and inhibitor of some ecto-ATPases. These two reagents inhibited the Mg2+-dependent ATPase activity in a dose-dependent manner. This ecto-ATPase was stimulated by more than 90% by 50 mM d -galactose. Since previous results showed that d -galactose exposed on the surface of host cells is involved with T. foetus adhesion, the Mg2+-dependent ecto-ATPase may be involved with cellular adhesion and possible pathogenicity.
P.a. Chate - One of the best experts on this subject based on the ideXlab platform.
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Hexagonal nanosized molybdenum diselenide thin film deposited at 333 K by chemical method
Solid State Sciences, 2015Co-Authors: D.j. Sathe, P.a. ChateAbstract:Abstract Molybdenum diselenide thin films have been deposited on to stainless steel and glass substrates by the chemical process, using ammonium Molybdate, Sodium selenosulphite as a precursor sources and citric acid was used as a complexing agent. The structural and optical properties of the deposited films have been studied using X-ray diffraction and optical absorption techniques, respectively. XRD studies reveal that the films are polycrystalline with hexagonal crystal structure. Optical absorption study shows the presence of direct transition with band gap energy 1.51 eV. EDAX analysis shows that the films are nearly stoichiometry of Mo: Se: 1:2. The configuration of fabricated cell is n -MoSe 2 | NaI (2 M) + I 2 (1 M) | C (graphite) yielded a conversion efficiency of 1.08%.
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A novel route for synthesis, characterization of molybdenum diselenide thin films and their photovoltaic applications
Journal of Materials Science: Materials in Electronics, 2013Co-Authors: D.j. Sathe, P.a. Chate, P. P. Hankare, A. H. Manikshete, A. S. AswarAbstract:Molybdenum diselenide thin films were deposited by chemical method. The precursor solution contains ammonium Molybdate, Sodium selenosulphite with hydrazine hydrate as a reducing agent. Various preparative conditions were optimized for the formation of thin films. The X-ray diffraction pattern shows that thin films have a layer-hexagonal phase. EDAX analysis shows that the films are nearly stoichiometric of Mo: Se: 1:2. Optical properties show a direct band gap nature with band gap energy 1.43 eV and having specific electrical conductivity in the order of 10^−5 (Ωcm)^−1. The configuration of fabricated cell is n -MoSe_2 | NaI (2 M) + I_2 (1 M) | C (graphite). The photoelectrochemical characterization of the films is carried out by studying current–voltage characterization, capacitance–voltage and power output characteristics. The fill factor and efficiency of the cell were found to be 34.22 and 1.01 % respectively.
D.j. Sathe - One of the best experts on this subject based on the ideXlab platform.
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Hexagonal nanosized molybdenum diselenide thin film deposited at 333 K by chemical method
Solid State Sciences, 2015Co-Authors: D.j. Sathe, P.a. ChateAbstract:Abstract Molybdenum diselenide thin films have been deposited on to stainless steel and glass substrates by the chemical process, using ammonium Molybdate, Sodium selenosulphite as a precursor sources and citric acid was used as a complexing agent. The structural and optical properties of the deposited films have been studied using X-ray diffraction and optical absorption techniques, respectively. XRD studies reveal that the films are polycrystalline with hexagonal crystal structure. Optical absorption study shows the presence of direct transition with band gap energy 1.51 eV. EDAX analysis shows that the films are nearly stoichiometry of Mo: Se: 1:2. The configuration of fabricated cell is n -MoSe 2 | NaI (2 M) + I 2 (1 M) | C (graphite) yielded a conversion efficiency of 1.08%.
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A novel route for synthesis, characterization of molybdenum diselenide thin films and their photovoltaic applications
Journal of Materials Science: Materials in Electronics, 2013Co-Authors: D.j. Sathe, P.a. Chate, P. P. Hankare, A. H. Manikshete, A. S. AswarAbstract:Molybdenum diselenide thin films were deposited by chemical method. The precursor solution contains ammonium Molybdate, Sodium selenosulphite with hydrazine hydrate as a reducing agent. Various preparative conditions were optimized for the formation of thin films. The X-ray diffraction pattern shows that thin films have a layer-hexagonal phase. EDAX analysis shows that the films are nearly stoichiometric of Mo: Se: 1:2. Optical properties show a direct band gap nature with band gap energy 1.43 eV and having specific electrical conductivity in the order of 10^−5 (Ωcm)^−1. The configuration of fabricated cell is n -MoSe_2 | NaI (2 M) + I_2 (1 M) | C (graphite). The photoelectrochemical characterization of the films is carried out by studying current–voltage characterization, capacitance–voltage and power output characteristics. The fill factor and efficiency of the cell were found to be 34.22 and 1.01 % respectively.