Artículo

Ulloa, R.M.; Muschietti, J.P.; Veron, M.; Torres, H.N.; Tellez-Iñón, M.T. "Purification and characterization of a soluble nucleoside diphosphate kinase in Trypanosoma cruzi" (1995) Molecular and Biochemical Parasitology. 70(1-2):119-129
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Abstract:

A soluble nucleoside diphosphate kinase (NDP kinase) was purified and characterized in epimastigote forms of Trypanosoma cruzi. The enzyme was purified by affinity chromatography on Blue-agarose and Q-Sepharose columns and by FPLC on a Superose 12 column. A membrane-associated NDP kinase was identified which accounts for 30% of total enzymatic activity. Western blot analysis of the soluble NDP kinase revealed a 16.5-kDa monomer recognized by polyclonal antibodies to NDP kinase from Dictyostelium discoideum, Candida albicans or human. Most of the T. cruzi NDP kinase is found in the cell as a hexamer composed of 16.5-kDa monomers. The Km values of the enzyme for ATP, GDP and dTDP were 0.2 ± 0.008 mM, 0.125 ± 0.012 mM and 0.4 ± 0.009 mM, respectively. The parasite enzyme was stable, remained active at 65°C and was found to tolerate up to 2.5 M urea. The 16.5-kDa subunit was phosphorylated with [γ-32P]ATP or thiophosphorylated with [35S]GTPγS. The incubation of the 32P-labelled phosphoenzyme with unlabelled nucleoside 5′-diphosphates resulted in the formation of 32P-labelled nucleoside 5′-triphosphates without strict base specificity, indicating that the reaction mechanism of the T. cruzi enzyme is the same as reported for other NDP kinases. When the phosphoenzyme was incubated with a mixture of nucleoside 5′-diphosphates, GTP was preferentially formed. © 1995.

Registro:

Documento: Artículo
Título:Purification and characterization of a soluble nucleoside diphosphate kinase in Trypanosoma cruzi
Autor:Ulloa, R.M.; Muschietti, J.P.; Veron, M.; Torres, H.N.; Tellez-Iñón, M.T.
Filiación:Instituto de Investigaciones en Ingeniería Genética y Biología Molecular, Facultad de Ciencias Exactas y Naturales. Obligado 2490, 1428 Buenos Aires, Argentina
Unité de Biochimie Cellulaire, Institut Pasteur, Paris, France
Palabras clave:Nucleoside diphosphate kinase; Phosphoenzyme intermediate; Trypanosoma cruzi; nucleoside diphosphate kinase; article; controlled study; enzyme analysis; enzyme purification; immunoblotting; nonhuman; priority journal; trypanosoma cruzi; Adenosine Triphosphate; Animal; Cross Reactions; Guanosine Diphosphate; Kinetics; Nucleoside-Diphosphate Kinase; Phosphorylation; Protein Conformation; Protein Denaturation; Protozoan Proteins; Solubility; Species Specificity; Substrate Specificity; Support, Non-U.S. Gov't; Temperature; Thymine Nucleotides; Trypanosoma cruzi; Candida albicans; Dictyostelium discoideum; Trypanosoma; Trypanosoma cruzi
Año:1995
Volumen:70
Número:1-2
Página de inicio:119
Página de fin:129
DOI: http://dx.doi.org/10.1016/0166-6851(95)00016-T
Título revista:Molecular and Biochemical Parasitology
Título revista abreviado:Mol. Biochem. Parasitol.
ISSN:01666851
CODEN:MBIPD
CAS:Adenosine Triphosphate, 56-65-5; Guanosine Diphosphate, 146-91-8; Nucleoside-Diphosphate Kinase, EC 2.7.4.6; Protozoan Proteins; thymidine 3',5'-diphosphate, 2863-04-9; Thymine Nucleotides
Registro:https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_01666851_v70_n1-2_p119_Ulloa

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Citas:

---------- APA ----------
Ulloa, R.M., Muschietti, J.P., Veron, M., Torres, H.N. & Tellez-Iñón, M.T. (1995) . Purification and characterization of a soluble nucleoside diphosphate kinase in Trypanosoma cruzi. Molecular and Biochemical Parasitology, 70(1-2), 119-129.
http://dx.doi.org/10.1016/0166-6851(95)00016-T
---------- CHICAGO ----------
Ulloa, R.M., Muschietti, J.P., Veron, M., Torres, H.N., Tellez-Iñón, M.T. "Purification and characterization of a soluble nucleoside diphosphate kinase in Trypanosoma cruzi" . Molecular and Biochemical Parasitology 70, no. 1-2 (1995) : 119-129.
http://dx.doi.org/10.1016/0166-6851(95)00016-T
---------- MLA ----------
Ulloa, R.M., Muschietti, J.P., Veron, M., Torres, H.N., Tellez-Iñón, M.T. "Purification and characterization of a soluble nucleoside diphosphate kinase in Trypanosoma cruzi" . Molecular and Biochemical Parasitology, vol. 70, no. 1-2, 1995, pp. 119-129.
http://dx.doi.org/10.1016/0166-6851(95)00016-T
---------- VANCOUVER ----------
Ulloa, R.M., Muschietti, J.P., Veron, M., Torres, H.N., Tellez-Iñón, M.T. Purification and characterization of a soluble nucleoside diphosphate kinase in Trypanosoma cruzi. Mol. Biochem. Parasitol. 1995;70(1-2):119-129.
http://dx.doi.org/10.1016/0166-6851(95)00016-T