title: Kinetic and biochemical characterization of Trypanosoma evansi nucleoside triphosphate diphosphohydrolase.
abstract: Nucleoside triphosphate diphospho-hydrolases (NTPDases) catalyze the hydrolysis of several nucleosides tri and diphosphate playing major roles in eukaryotes including purinergic signaling, inflammation, hemostasis, purine salvage and host-pathogen interactions. These enzymes have been recently described in parasites where several evidences indicated their involvement in virulence and infection. Here, we have investigated the presence of NTPDase in the genome of Trypanosoma evansi. Based on the genomic sequence from Trypanosoma brucei, we have amplified an 1812 gene fragment corresponding to the T. evansi NTPDase gene. The protein was expressed in the soluble form and purified to homogeneity and enzymatic assays were performed confirming the enzyme identity. Kinetic parameters and substrate specificity were determined. The dependence of cations on enzymatic activity was investigated indicating the enzyme is stimulated by divalent cations and carbohydrates but inhibited by sodium. Bioinformatic analysis indicates the enzyme is a membrane bound protein facing the extracellular side of the cell with 98% identity to the T. brucei homologous NTPDase gene.
authors.affiliation: Departamento de Produção Animal e Alimentos, Universidade do Estado de Santa Catarina, Av. Luiz de Camões, 2090 Lages, Santa Catarina, Brazil.
authors.forename: Paulo Henrique Exterchoter
authors.initials: PH
authors.lastname: Weiss
authors.affiliation: Departamento de Produção Animal e Alimentos, Universidade do Estado de Santa Catarina, Av. Luiz de Camões, 2090 Lages, Santa Catarina, Brazil.
authors.forename: Franciane
authors.initials: F
authors.lastname: Batista
authors.affiliation: Área das Ciências Biológicas e da Saúde, Universidade do Oeste de Santa Catarina, Av. Getúlio Vargas, 2125 Joaçaba, Santa Catarina, Brazil.
authors.forename: Glauber
authors.initials: G
authors.lastname: Wagner
authors.affiliation: Departamento de Produção Animal e Alimentos, Universidade do Estado de Santa Catarina, Av. Luiz de Camões, 2090 Lages, Santa Catarina, Brazil.
authors.forename: Maria de Lourdes Borba
authors.initials: Mde L
authors.lastname: Magalhães
authors.affiliation: Departamento de Produção Animal e Alimentos, Universidade do Estado de Santa Catarina, Av. Luiz de Camões, 2090 Lages, Santa Catarina, Brazil. Electronic address: luiz.miletti@udesc.br.
authors.forename: Luiz Claudio
authors.initials: LC
authors.lastname: Miletti
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D004795:Enzyme Stability; D005260:Female; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D043583:Nucleoside-Triphosphatase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D015800:Protozoan Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D016415:Sequence Alignment; D013379:Substrate Specificity; D013696:Temperature; D014345:Trypanosoma / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D014352:Trypanosomiasis / Q000469:parasitology*
chemical_list: D015800:Protozoan Proteins; D043583:Nucleoside-Triphosphatase
keywords: NTPDase; Steady-state kinetics; Trypanosoma evansi