| Author | Morgado-Díaz, José Andrés | |
| Author | Lopez, Raquel Elisa da Silva | |
| Author | Alves, Carlos Roberto | |
| Author | Soares, Maurilio José | |
| Author | Corte-Real, Suzana | |
| Author | De Simone, Salvatore Giovanni | |
| Access date | 2019-11-12T12:44:11Z | |
| Available date | 2019-11-12T12:44:11Z | |
| Document date | 2005 | |
| Citation | MORGADO-DIAZ, José Andrés et al. Subcellular localization of an intracellular serine protease of 68 kDa in Leishmania (Leishmania) amazonensis promastigotes. Memórias do Insituto Oswaldo Cruz, Rio de Janeiro, v. 100, n. 4, p. 377-383, July 2005. | pt_BR |
| ISSN | 0074-0276 | pt_BR |
| URI | https://www.arca.fiocruz.br/handle/icict/36996 | |
| Language | eng | pt_BR |
| Publisher | Fundação Oswaldo Cruz. Instituto Oswaldo Cruz | pt_BR |
| Rights | open access | |
| Subject in Portuguese | Leishmania | pt_BR |
| Subject in Portuguese | Protease serina | pt_BR |
| Subject in Portuguese | Localização subcelular | pt_BR |
| Subject in Portuguese | Tripanosomatídeo | pt_BR |
| Title | Subcellular localization of an intracellular serine protease of 68 kDa in Leishmania (Leishmania) amazonensis promastigotes | pt_BR |
| Type | Article | |
| DOI | 10.1590/s0074-02762005000400007 | |
| Abstract | Here we report the subcellular localization of an intracellular serine protease of 68 kDa in axenic promastigotes of Leishmania (Leishmania) amazonensis, using subcellular fractionation, enzymatic assays, immunoblotting, and immunocytochemistry. All fractions were evaluated by transmission electron microscopy and the serine protease activity was measured during the cell fractionation procedure using alpha-N-r-tosyl-L-arginine methyl ester (L-TAME) as substrate, phenylmethylsulphone fluoride (PMSF) and L-1-tosylamino-2-phenylethylchloromethylketone (TPCK) as specific inhibitors. The enzymatic activity was detected mainly in a membranous vesicular fraction (6.5-fold enrichment relative to the whole homogenate), but also in a crude plasma membrane fraction (2.0-fold). Analysis by SDS-PAGE gelatin under reducing conditions demonstrated that the major proteolytic activity was found in a 68 kDa protein in all fractions studied. A protein with identical molecular weight was also recognized in immunoblots by a polyclonal antibody against serine protease (anti-SP), with higher immunoreactivity in the vesicular fraction. Electron microscopic immunolocalization using the same polyclonal antibody showed the enzyme present at the cell surface, as well as in cytoplasmic membranous compartments of the parasite. Our findings indicate that the internal location of this serine protease in L. amazonensis is mainly restricted to the membranes of intracellular compartments resembling endocytic/exocytic elements. | pt_BR |
| Affilliation | Instituto Nacional de Câncer. Centro de Pesquisas. Divisão de Biologia Celular. Grupo de Biologia Estrutural. Rio e Janeiro, RJ, Brasil. | pt_BR |
| Affilliation | Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Departamento de Bioquímica e Biologia Molecular. Laboratório de Bioquímica de Proteínas e Peptídios. Rio de Janeiro, RJ, Brasil. | pt_BR |
| Affilliation | Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Departamento de Bioquímica e Biologia Molecular. Laboratório de Bioquímica de Proteínas e Peptídios. Rio de Janeiro, RJ, Brasil. | pt_BR |
| Affilliation | Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Departamento de Ultra-estrutura e Biologia Celular. Rio de Janeiro, RJ, Brasil. | pt_BR |
| Affilliation | Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Departamento de Ultra-estrutura e Biologia Celular. Rio de Janeiro, RJ, Brasil. | pt_BR |
| Affilliation | Universidade Federal Fluminense. Instituto de Biologia. Departamento de Biologia Celular e Molecular. Niterói, RJ, Brasil. | pt_BR |
| Subject | Leishmania | pt_BR |
| Subject | Serine protease | pt_BR |
| Subject | Subcellular localization | pt_BR |
| Subject | Trypanosomatid | pt_BR |
| e-ISSN | 1678-8060 | |
