Artículos de revistas
Trypanosoma Cruzi: Isolation And Characterization Of A Proteinase
Registro en:
Experimental Parasitology. , v. 52, n. 2, p. 199 - 209, 1981.
144894
10.1016/0014-4894(81)90075-8
2-s2.0-0019833366
Autor
Rangel H.A.
Araujo P.M.F.
Repka D.
Costa M.G.
Institución
Resumen
Lysates of Trypanosoma cruzi epimastigotes were able to hydrolyze casein (Km = 2.5 mg/ml) as well as bovine and human hemoglobins (Km = 12.2 mg/ml); there was optimum activity was around pH 7.0. The proteinase activity detected with these substrates was enhanced by sodium diaminotetraacetate (EDTA) and reducing agents (SO2- 3, mercaptoethanol, cysteine) and was inhibited by sulfhydryl reagents, thus suggesting an SH-dependent enzyme. Purification (60×) of the proteinase was carried out as follows: (1) precipitation at -20 C, pH 4.5, with 80% acetone, (2) gel filtration on Sephadex G-200, (3) affinity chromatography on Sepharose 4B covalently linked to p-aminophenyl mercuric acetate. Only a single component (with an estimated molecular weight of 60,000) was detected in purified preparations by polyacrylamide gel electrophoresis. However, in addition to the major component identified as a proteinase, crossed immunoelectrophoresis experiments indicated the presence of at least three other antigens that apparently were devoid of proteinase activity. Optimum pH activity of the purified preparations was around pH 6.0 for casein and pH 3.0 for hemoglobins, but these activities probably are due to the one enzyme since they were altered identically by the same agents. © 1981. 52 2 199 209 Adair, Robinson, The specific refraction increments of serum albumin and serum globin (1930) Biochemical Journal, 24, pp. 933-1011 Anson, The estimation of pepsin trypsin papain and cathepsin with hemoglobin (1938) The Journal of General Physiology, 22, pp. 79-89 Bongertz, Hungerer, Trypanosoma cruzi: Isolation and characterization of a protease (1978) Experimental Parasitology, 45, pp. 8-18 Dimroth, Ueber die Mercurrirung aromatischer Verbindungen (1902) Berichte der deutschen chemischen Gesellschaft, 35, pp. 2032-2045 Dubois, Gilles, Hamilton, Rebers, Smith, Colorimetric method for determination of sugars and related substances (1956) Analytical Chemistry, 28, pp. 350-356 Fairbanks, Steck, Wallach, Electrophoretic analysis of the major polypeptides of the human erythrocyte membrane (1971) Biochemistry, 10, pp. 2606-2617 Fernandes, Castelani, Growth characteristics and chemical composition of Trypanosoma cruzi (1966) Experimental Parasitology, 18, pp. 195-202 Flodin, (1962) Dextran Gels and Their Applications in Gel Filtration, , 3rd ed., Phamacia, Uppsala Holzer, Betz, Ebner, Intracellular proteinase in microorganisms (1975) Current Topics in Cellular Regulation, 9, pp. 103-156. , B.L. Horecker, E.R. Stadtman, Adacemic Press, New York Itow, Camargo, Proteolytic activities in cell extracts of Trypanosoma cruzi (1977) Journal of Protozoology, 24, pp. 591-595 Kabat, Mayer, (1961) Experimental Immunochemistry, , 2nd ed., Charles C. Thomas, Springfield, Ill Lineweaver, Burke, The determination of the enzyme dissociation constants (1934) Journal of the American Chemical Society, 56, pp. 658-666 Lowry, Rosebrough, Farr, Randall, Protein measurement with the Folin phenol reagent (1951) Journal of Biological Chemistry, 193, pp. 265-275 Means, Feeney, (1971) Chemical Modification of Proteins, , Holden-Day, San Francisco O'Daly, A new liquid medium for Trypanosoma cruzi (1975) Journal of Protozoology, 22, pp. 265-270 O'Daly, Effect of fetal calf serum fractions and proteins on division and transformation of Trypanosoma cruzi in vitro (1976) Journal of Protozoology, 23, pp. 577-583 Ouchterlony, Diffusion-in-gel methods for immunological analysis (1958) Progress in Allergy, 5, pp. 1-78 Rangel, Repka, Costa, Proteolytic activity of Trypanosoma cruzi crude extracts (1976) Resumos da IV Reunião Anual sobre Pesquisa Básica em Doença de Chagas, p. 30. , Caxambú-Minas Gerais, Brasil, 27–29 De Outubro Rangel, Repka, Araujo, Costa, A neutral proteinase of the epimastigote forms of Trypanosoma cruzi (1977) Resumos da IV Reunião Anual sobre Pesquisa Básica em Doença de Chagas, p. 58. , Caxambú-Minas Gerais, Brasil, 5–7 de Novembro Repka, Rangel, Costa, Atividade proteolĭtica dos extratos de Trypanosoma cruzi (1972) Resumos da XXIV reunião Anual da Sociedade Brasileira para o Progresso da Ciência, 24, p. 296. , São Paulo de 2 a 8 de Julho Silva, Nussenzweig, Sobre uma cepa de Trypanosoma cruzi altamente virulente para o camundomin o branco (1953) Folia Clinica et Biologica São Paulo, 20, pp. 191-208 Sluyterman, Wijdenes, An agarose mercurial column for the separation of mercaptopapain and nonmercaptopapain (1970) Biochimica et Biophysica Acta (BBA) - Protein Structure, 200, pp. 590-593 Uriel, (1971) Methods in Immunology and Immumochemistry, pp. 294-321. , C.A. Williams, M.W. Chase, Academic Press, New York Walsh, Wilcox, Serine proteases (1970) Methods in Enzymology, 19, pp. 31-41. , G.E. Perlmann, L. Laszlo, Academic Press, New York Weeke, Crossed immunoelectrophoresis (1973) A Manual of Quantitative Immunoelectrophoresis: Methods and Applications, pp. 47-56. , N.H. Axelsen, J. Kroll, B. Weeke, Universitesforlaget, Oslo