The major leucyl aminopeptidase of Trypanosoma cruzi (LAPTc) assembles into a homohexamer and belongs to the M17 family of metallopeptidases

<p>Abstract</p> <p>Background</p> <p>Pathogens depend on peptidase activities to accomplish many physiological processes, including interaction with their hosts, highlighting parasitic peptidases as potential drug targets. In this study, a major leucyl aminopeptidolytic activity was identified in <it>Trypanosoma cruzi</it>, the aetiological agent of Chagas disease.</p> <p>Results</p> <p>The enzyme was isolated from epimastigote forms of the parasite by a two-step chromatographic procedure and associated with a single 330-kDa homohexameric protein as determined by sedimentation velocity and light scattering experiments. Peptide mass fingerprinting identified the enzyme as the predicted <it>T. cruzi </it>aminopeptidase EAN97960. Molecular and enzymatic analysis indicated that this leucyl aminopeptidase of <it>T. cruzi </it>(LAPTc) belongs to the peptidase family M17 or leucyl aminopeptidase family. LAPTc has a strong dependence on neutral pH, is mesophilic and retains its oligomeric form up to 80°C. Conversely, its recombinant form is thermophilic and requires alkaline pH.</p> <p>Conclusions</p> <p>LAPTc is a 330-kDa homohexameric metalloaminopeptidase expressed by all <it>T. cruzi </it>forms and mediates the major parasite leucyl aminopeptidolytic activity. Since biosynthetic pathways for essential amino acids, including leucine, are lacking in <it>T. cruzi</it>, LAPTc could have a function in nutritional supply.</p

Anti-IL-2 Treatment Impairs the Expansion of Treg Cell Population during Acute Malaria and Enhances the Th1 Cell Response at the Chronic Disease

Plasmodium chabaudi infection induces a rapid and intense splenic CD4+ T cell response that contributes to both disease pathogenesis and the control of acute parasitemia. The subsequent development of clinical immunity to disease occurs concomitantly with the persistence of low levels of chronic parasitemia. The suppressive activity of regulatory T (Treg) cells has been implicated in both development of clinical immunity and parasite persistence. To evaluate whether IL-2 is required to induce and to sustain the suppressive activity of Treg cells in malaria, we examined in detail the effects of anti-IL-2 treatment with JES6-1 monoclonal antibody (mAb) on the splenic CD4+ T cell response during acute and chronic P. chabaudi AS infection in C57BL/6 mice. JES6-1 treatment on days 0, 2 and 4 of infection partially inhibits the expansion of the CD4+CD25+Foxp3+ cell population during acute malaria. Despite the concomitant secretion of IL-2 and expression of high affinity IL-2 receptor by large CD4+ T cells, JES6-1 treatment does not impair effector CD4+ T cell activation and IFN-γ production. However, at the chronic phase of the disease, an enhancement of cellular and humoral responses occurs in JES6-1-treated mice, with increased production of TNF-α and parasite-specific IgG2a antibodies. Furthermore, JES6-1 mAb completely blocked the in vitro proliferation of CD4+ T cells from non-treated chronic mice, while it further increased the response of CD4+ T cells from JES6-1-treated chronic mice. We conclude that JES6-1 treatment impairs the expansion of Treg cell population during early P. chabaudi malaria and enhances the Th1 cell response in the late phase of the disease

The Trypanosoma cruzi Virulence Factor Oligopeptidase B (OPBTc) Assembles into an Active and Stable Dimer

Oligopeptidase B, a processing enzyme of the prolyl oligopeptidase family, is considered as an important virulence factor in trypanosomiasis. Trypanosoma cruzi oligopeptidase B (OPBTc) is involved in host cell invasion by generating a Ca2+-agonist necessary for recruitment and fusion of host lysosomes at the site of parasite attachment. The underlying mechanism remains unknown and further structural and functional characterization of OPBTc may help clarify its physiological function and lead to the development of new therapeutic molecules to treat Chagas disease. In the present work, size exclusion chromatography and analytical ultracentrifugation experiments demonstrate that OPBTc is a dimer in solution, an association salt and pH-resistant and independent of intermolecular disulfide bonds. The enzyme retains its dimeric structure and is fully active up to 42°C. OPBTc is inactivated and its tertiary, but not secondary, structure is disrupted at higher temperatures, as monitored by circular dichroism and fluorescence spectroscopy. It has a highly stable secondary structure over a broad range of pH, undergoes subtle tertiary structure changes at low pH and is less stable under moderate ionic strength conditions. These results bring new insights into the structural properties of OPBTc, contributing to future studies on the rational design of OPBTc inhibitors as a promising strategy for Chagas disease chemotherapy

Injeção de cama de aviário no sulco de plantio.

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Nicotinamide and Azospirillum brasilense improves the quality of Coffea arabica seedlings

ABSTRACT The use of biostimulants in coffee seedlings can promote gains in their growth and quality. Thus, this study aimed to evaluate the growth and quality characteristics of Coffea arabica seedlings under the effect of the nicotinamide and Azospirillum brasilense application. The experimental design was randomized blocks with treatments arranged in a 5 × 2 factorial scheme, with four replicates. The treatments resulted from the use of five doses of nicotinamide (0, 30, 60, 90, and 120 mg L-1 of water) combined with the absence and presence of Azospirillum brasilense applied to Coffea arabica seedlings from Catuaí Vermelho 144 cultivar. Plant height, stem diameter, number of leaves, leaf area, shoot dry mass, root dry mass, plant height:stem diameter ratio, shoot:root dry mass ratio, plant height:shoot dry mass ratio, and Dickson quality index were evaluated. The combined or isolated use of A. brasilense and nicotinamide, up to a dose of 33.5 mg L-1, increased the biometric characteristics and dry mass accumulation. However, using nicotinamide doses between 30 and 61.8 g L-1 increased the quality of Coffea arabica seedlings. The synergistic effect of the use of A. brasilense and nicotinamide was verified for the growth and quality of Coffea arabica seedlings.</div