Hostname: page-component-cd9895bd7-gbm5v Total loading time: 0 Render date: 2024-12-29T03:00:39.835Z Has data issue: false hasContentIssue false

A zymographic study of metalloprotease activities in extracts and extracellular secretions of Leishmania (Viannia) braziliensis strains

Published online by Cambridge University Press:  03 October 2005

P. CUERVO
Affiliation:
Departamento de Medicina Tropical, Instituto Oswaldo Cruz, FIOCRUZ, Rio de Janeiro, Brasil Centro Internacional de Entrenamiento e Investigaciones Médicas CIDEIM, Cali, Colombia Departamento de Imunologia, Instituto Oswaldo Cruz, FIOCRUZ, Rio de Janeiro, Brasil
L. SABÓIA-VAHIA
Affiliation:
Departamento de Bioquímica e Biologia Molecular, Instituto Oswaldo Cruz, FIOCRUZ, Rio de Janeiro, Brasil Instituto de Biofisica, Carlos Chagas Filho, UFRJ, Rio de Janeiro, Brasil
F. COSTA SILVA-FILHO
Affiliation:
Instituto de Biofisica, Carlos Chagas Filho, UFRJ, Rio de Janeiro, Brasil
O. FERNANDES
Affiliation:
Departamento de Medicina Tropical, Instituto Oswaldo Cruz, FIOCRUZ, Rio de Janeiro, Brasil
E. CUPOLILLO
Affiliation:
Departamento de Imunologia, Instituto Oswaldo Cruz, FIOCRUZ, Rio de Janeiro, Brasil
J. B. DE JESUS
Affiliation:
Departamento de Bioquímica e Biologia Molecular, Instituto Oswaldo Cruz, FIOCRUZ, Rio de Janeiro, Brasil Instituto de Biofisica, Carlos Chagas Filho, UFRJ, Rio de Janeiro, Brasil

Abstract

Proteolytic activities of 5 strains of Leishmania (Viannia) braziliensis isolated from Brazilian and Colombian patients, presenting distinct clinical manifestations, were characterized and compared using whole-promastigote extracts and extracellular secretions. Zymographic assays concerning whole-cell extracts and supernatants resulted in the detection of high molecular weight bands, ranging from 50 to 125 kDa. Proteolytic activities from both whole-cell extracts and supernatants were optimal in a pH range 5·5 to 9·0 for all analysed strains. Such protease activities were inhibited when 10 mM 1,10-phenanthroline was assayed, strongly suggesting that the enzymes responsible for hydrolysis of the substrate belong to the metalloproteases class. Distinct profiles of metalloproteases were observed among the studied L. (V.) braziliensis strains. Differences among the microorganisms might be related to the geographical origin of the strains and/or to the clinical presentation.

Type
Research Article
Copyright
2005 Cambridge University Press

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

REFERENCES

Alexander, J., Coombs, G. H. and Mottram, J. C. ( 1998). Leishmania mexicana cysteine proteinase-deficient mutants have attenuated virulence for mice and potentiate a Th1 response. Journal of Immunology 161, 67946801.Google Scholar
Alves, C. R., Corte-Real, S., Bourguignon, S. C., Chaves, C. S. and Saraiva, E. M. ( 2005). Leishmania amazonensis: early proteinase activities during promastigote-amastigote differentiation in vitro. Experimental Parasitology 109, 3848.CrossRefGoogle Scholar
Bates, P. A., Hermes, I. and Dwyer, D. M. ( 1989). Leishmania donovani: immunochemical localization and secretory mechanism of soluble acid phosphatase. Experimental Parasitology 68, 335346.CrossRefGoogle Scholar
Beynon, R. and Bond, J. S. ( 2001). Proteolytic Enzymes, 2nd Edn. Oxford University Press, New York.
Blum, J. J. and Opperdoes, F. R. ( 1994). Secretion of sucrase by Leishmania donovani. Journal of Eukaryotic Microbiology 41, 228231.CrossRefGoogle Scholar
Bontempi, E. and Cazzulo, J. J. ( 1990). Digestion of human immunoglobulin G by the major cysteine proteinase (cruzipain) from Trypanosoma cruzi. FEMS Microbiology Letters 58, 337341.CrossRefGoogle Scholar
Bouvier, J., Etges, R. J. and Bordier, C. ( 1985). Identification and purification of membrane and soluble forms of the major surface protein of Leishmania promastigotes. The Journal of Biological Chemistry 260, 1550415509.Google Scholar
Brittingham, A., Morrison, C. J., McMaster, W. R., McGwire, B. S., Chang, K. P. and Mosser, D. M. ( 1995). Role of the Leishmania surface protease gp63 in complement fixation, cell adhesion, and resistance to complement-mediated lysis. Journal of Immunology 155, 31023111.CrossRefGoogle Scholar
Brittingham, A., Chen, G., McGwire, B. S., Chang, K. P. and Mosser, D. M. ( 1999). Interaction of Leishmania gp63 with cellular receptors for fibronectin. Infection and Immunity 67, 44774484.Google Scholar
Buxbaum, L. U., Denise, H., Coombs, G. H., Alexander, J., Mottram, J. C. and Scott, P. ( 2003). Cysteine protease B of Leishmania mexicana inhibits host Th1 responses and protective immunity. Journal of Immunology 171, 37113717.CrossRefGoogle Scholar
Canto-Lara, S. B., Cardenas-Maruffo, M. F., Vargas-Gonzalez, A. and Andrade-Narvaez, F. ( 1998). Isoenzyme characterization of Leishmania isolated from human cases with localized cutaneous leishmaniasis from the State of Campeche, Yucatan Peninsula, Mexico. American Journal of Tropical Medicine and Hygiene 58, 444447.CrossRefGoogle Scholar
Chang, C. S. and Chang, K. P. ( 1986). Monoclonal antibody affinity purification of a Leishmania membrane glycoprotein and its inhibition of leishmania-macrophage binding. Proceedings of the National Academy of Sciences, USA 83, 100104.CrossRefGoogle Scholar
Chaudhuri, G., Chaudhuri, M., Pan, A. and Chang, K. P. ( 1989). Surface acid proteinase (gp63) of Leishmania mexicana. A metalloenzyme capable of protecting liposome-encapsulated proteins from phagolysosomal degradation by macrophages. The Journal of Biological Chemistry 264, 74837489.Google Scholar
Colomer-Gould, V., Glvao Quintao, L., Keithly, J. and Nogueira, N. ( 1985). A common major surface antigen on amastigotes and promastigotes of Leishmania species. The Journal of Experimental Medicine 162, 902916.CrossRefGoogle Scholar
Coombs, G. H. ( 1982). Proteinases of Leishmania mexicana and other flagellate protozoa. Parasitology 84, 149155.CrossRefGoogle Scholar
Coppi, A., Pinzon-Ortiz, C., Hutter, C. and Sinnis, P. ( 2005). The Plasmodium circumsporozoite protein is proteolytically processed during cell invasion. The Journal of Experimental Medicine 201, 2733.CrossRefGoogle Scholar
Cupolillo, E., Brahim, L. R., Toaldo, C. B., de Oliveira-Neto, M. P., de Brito, M. E., Falqueto, A., de Farias Naiff, M. and Grimaldi, G. Jr. ( 2003). Genetic polymorphism and molecular epidemiology of Leishmania (Viannia) braziliensis from different hosts and geographic areas in Brazil. Journal of Clinical Microbiology 41, 31263132.CrossRefGoogle Scholar
De Araujo Soares, R. M., dos Santos, A. L., Bonaldo, M. C., de Andrade, A. F., Alviano, C. S., Angluster, J. and Goldenberg, S. ( 2003). Leishmania (Leishmania) amazonensis: differential expression of proteinases and cell-surface polypeptides in avirulent and virulent promastigotes. Experimental Parasitology 104, 104112.CrossRefGoogle Scholar
Etges, R. ( 1992). Identification of a surface metalloproteinase on 13 species of Leishmania isolated from humans, Crithidia fasciculata, and Herpetomonas samuelpessoai. Acta Tropica 50, 205217.CrossRefGoogle Scholar
Etges, R., Bouvier, J. and Bordier, C. ( 1986). The major surface protein of Leishmania promastigotes is a protease. The Journal of Biological Chemistry 261, 90989101.Google Scholar
Etges, R. J., Bouvier, J., Hoffman, R. and Bordier, C. ( 1985). Evidence that the major surface proteins of three Leishmania species are structurally related. Molecular and Biochemical Parasitology 14, 141149.CrossRefGoogle Scholar
Fong, D. and Chang, K. P. ( 1981). Tubulin biosynthesis in the developmental cycle of a parasitic protozoan, Leishmania mexicana: changes during differentiation of motile and nonmotile stages. Proceedings of the National Academy of Sciences, USA 78, 76247628.CrossRefGoogle Scholar
Gardiner, P. R., Jaffe, C. L. and Dwyer, D. M. ( 1984). Identification of cross-reactive promastigote cell surface antigens of some leishmanial stocks by 125I labeling and immunoprecipitation. Infection and Immunity 43, 637643.Google Scholar
Grimaldi, G. Jr. and Tesh, R. B. ( 1993). Leishmaniases of the New World: current concepts and implications for future research. Clinical Microbiology Reviews 6, 230250.CrossRefGoogle Scholar
Grimaldi, G. Jr., Tesh, R. B. and McMahon-Pratt, D. ( 1989). A review of the geographic distribution and epidemiology of leishmaniasis in the New World. American Journal of Tropical Medicine and Hygiene 41, 687725.CrossRefGoogle Scholar
Hajmova, M., Chang, K. P., Kolli, B. and Volf, P. ( 2004). Down-regulation of gp63 in Leishmania amazonensis reduces its early development in Lutzomyia longipalpis. Microbes and Infection/Institut Pasteur 6, 646649.CrossRefGoogle Scholar
Jacobson, R. L. and Schlein, Y. ( 1997). Cellulase activity of Leishmania major in the sandfly vector and in culture. Journal of Eukaryotic Microbiology 44, 216219.CrossRefGoogle Scholar
Jaffe, C. L. and Dwyer, D. M. ( 2003). Extracellular release of the surface metalloprotease, gp63, from Leishmania and insect trypanosomatids. Parasitology Research 91, 229237.CrossRefGoogle Scholar
Klemba, M. and Goldberg, D. E. ( 2002). Biological roles of proteases in parasitic protozoa. Annual Review of Biochemistry 71, 275305.CrossRefGoogle Scholar
Leon, L. L., Temporal, R. M., Soares, M. J. and Grimaldi, G. Jr. ( 1994). Proteinase activities during temperature-induced stage differentiation of species complexes of Leishmania. Acta Tropica 56, 289298.CrossRefGoogle Scholar
Liu, X. and Chang, K. P. ( 1992). Extrachromosomal genetic complementation of surface metalloproteinase (gp63)-deficient Leishmania increases their binding to macrophages. Proceedings of the National Academy of Sciences, USA 89, 49914995.CrossRefGoogle Scholar
Lockwood, B. C., North, M. J., Scott, K. I., Bremner, A. F. and Coombs, G. H. ( 1987). The use of a highly sensitive electrophoretic method to compare the proteinases of trichomonads. Molecular and Biochemical Parasitology 24, 8995.CrossRefGoogle Scholar
Mahmoudzadeh-Niknam, H. and McKerrow, J. H. ( 2004). Leishmania tropica: cysteine proteases are essential for growth and pathogenicity. Experimental Parasitology 106, 158163.CrossRefGoogle Scholar
Marsden, P. D. ( 1986). Mucosal leishmaniasis (“espundia” Escomel, 1911). Transactions of the Royal Society of Tropical Medicine and Hygiene 80, 859876.CrossRefGoogle Scholar
McGrath, M. E., Eakin, A. E., Engel, J. C., McKerrow, J. H., Craik, C. S. and Fletterick, R. J. ( 1995). The crystal structure of cruzain: a therapeutic target for Chagas' disease. Journal of Molecular Biology 247, 251259.CrossRefGoogle Scholar
McGwire, B. S., O'Connell, W. A., Chang, K. P. and Engman, D. M. ( 2002). Extracellular release of the glycosylphosphatidylinositol (GPI)-linked Leishmania surface metalloprotease, gp63, is independent of GPI phospholipolysis: implications for parasite virulence. The Journal of Biological Chemistry 277, 88028809.CrossRefGoogle Scholar
McGwire, B. S., Chang, K. P. and Engman, D. M. ( 2003). Migration through the extracellular matrix by the parasitic protozoan Leishmania is enhanced by surface metalloprotease gp63. Infection and Immunity 71, 10081010.CrossRefGoogle Scholar
Medina-Acosta, E., Karess, R. E. and Russell, D. G. ( 1993). Structurally distinct genes for the surface protease of Leishmania mexicana are developmentally regulated. Molecular and Biochemical Parasitology 57, 3145.CrossRefGoogle Scholar
Medina-Acosta, E., Karess, R. E., Schwartz, H. and Russell, D. G. ( 1989). The promastigote surface protease (gp63) of Leishmania is expressed but differentially processed and localized in the amastigote stage. Molecular and Biochemical Parasitology 37, 263273.CrossRefGoogle Scholar
Melo-Braga, M. B., da Rocha-Azevedo, B. and Silva-Filho, F. C. ( 2003). Tritrichomonas foetus: the role played by iron during parasite interaction with epithelial cells. Experimental Parasitology 105, 111120.CrossRefGoogle Scholar
Mottram, J. C., Brooks, D. R. and Coombs, G. H. ( 1998). Roles of cysteine proteinases of trypanosomes and Leishmania in host-parasite interactions. Current Opinion in Microbiology 1, 455460.CrossRefGoogle Scholar
Mottram, J. C., Souza, A. E., Hutchison, J. E., Carter, R., Frame, M. J. and Coombs, G. H. ( 1996). Evidence from disruption of the lmcpb gene array of Leishmania mexicana that cysteine proteinases are virulence factors. Proceedings of the National Academy of Sciences, USA 93, 60086013.CrossRefGoogle Scholar
North, M. J. and Coombs, G. H. ( 1981). Proteinases of Leishmania mexicana amastigotes and promastigotes: analysis by gel electrophoresis. Molecular and Biochemical Parasitology 3, 293300.CrossRefGoogle Scholar
Pupkis, M. F. and Coombs, G. H. ( 1984). Purification and characterization of proteolytic enzymes of Leishmania mexicana mexicana amastigotes and promastigotes. Journal of General Microbiology 130, 23752383.CrossRefGoogle Scholar
Que, X. and Reed, S. L. ( 2000). Cysteine proteinases and the pathogenesis of amebiasis. Clinical Microbiology Reviews 13, 196206.CrossRefGoogle Scholar
Robertson, C. D. and Coombs, G. H. ( 1992). Stage-specific proteinases of Leishmania mexicana mexicana promastigotes. FEMS Microbiology Letters 73, 127132.CrossRefGoogle Scholar
Rosenthal, P. J. ( 1999). Proteases of protozoan parasites. Advances in Parasitology 43, 105159.CrossRefGoogle Scholar
Russell, D. G. and Wilhelm, H. ( 1986). The involvement of the major surface glycoprotein (gp63) of Leishmania promastigotes in attachment to macrophages. Journal of Immunology 136, 26132620.Google Scholar
Sajid, M. and McKerrow, J. H. ( 2002). Cysteine proteases of parasitic organisms. Molecular and Biochemical Parasitology 120, 121.CrossRefGoogle Scholar
Saravia, N. G., Holguin, A. F., McMahon-Pratt, D. and D'Alessandro, A. ( 1985). Mucocutaneous leishmaniasis in Colombia: Leishmania braziliensis subspecies diversity. American Journal of Tropical Medicine and Hygiene 34, 714720.CrossRefGoogle Scholar
Schlein, Y. ( 1993). Leishmania and sandflies: interactions in the life-cycle and transmission. Parasitology Today 9, 255258.CrossRefGoogle Scholar
Schneider, P. and Glaser, T. A. ( 1993). Characterization of two soluble metalloexopeptidases in the protozoan parasite Leishmania major. Molecular and Biochemical Parasitology 62, 223231.CrossRefGoogle Scholar
Silva-Lopez, R. E., Morgado-Diaz, J. A., Alves, C. R., Corte-Real, S. and Giovanni-De-Simone, S. ( 2004). Subcellular localization of an extracellular serine protease in Leishmania (Leishmania) amazonensis. Parasitology Research 93, 328331.CrossRefGoogle Scholar
Weigle, K. A., Saravia, N. G., de Davalos, M., Moreno, L. H. and D'Alessandro, A. ( 1986). Leishmania braziliensis from the Pacific coast region of Colombia: foci of transmission, clinical spectrum and isoenzyme phenotypes. American Journal of Tropical Medicine and Hygiene 35, 722731.CrossRefGoogle Scholar
Yao, C., Leidal, K. G., Brittingham, A., Tarr, D. E., Donelson, J. E. and Wilson, M. E. ( 2002). Biosynthesis of the major surface protease GP63 of Leishmania chagasi. Molecular and Biochemical Parasitology 121, 119128.CrossRefGoogle Scholar
Young, D. G., Morales, A., Kreutzer, R. D., Alexander, J. B., Corredor, A., Tesh, R. B., Ferro de Carrasquilla, C. and de Rodriguez, C. ( 1987). Isolation of Leishmania braziliensis (Kinetoplastida: Trypanosomatidae) from cryopreserved Colombian sand flies (Diptera: Psychodidae). Journal of Medical Entomology 24, 587589.CrossRefGoogle Scholar