Abstract
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.
Highlights
Chagas disease (American Trypanosomiasis) is a multisystemic illness resulting from the infection with the intracellular protozoan parasite Trypanosoma cruzi, which is able to invade a wide variety of mammalian cells
OPBTc is a Dimeric Enzyme The active recombinant OPBTc was expressed in E. coli and shares similar properties with its native form purified from T. cruzi [7,22,27]
We examined some structural properties of OPBTc, a key protease for triggering Ca2+-signaling in both parasite and host cells, a crucial step to T. cruzi infection process
Summary
Chagas disease (American Trypanosomiasis) is a multisystemic illness resulting from the infection with the intracellular protozoan parasite Trypanosoma cruzi, which is able to invade a wide variety of mammalian cells It affects millions of people in Latin America and represents a major public health problem because of its high rates of morbidity and mortality due to clinical complications at the chronic phase [1]. This scenario requires functional and molecular characterization of T. cruzi virulence factors that could serve as the basis for vaccine development or as targets for Chagas disease chemotherapy. Specific silencing of OPBTc gene greatly inhibited the infective capacity of trypomastigotes both in vitro and in vivo, revealing OPBTc as a T. cruzi virulence factor and, a good target for developing new drugs to treat T. cruzi infections [7,8]
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