Details

Autor(en) / Beteiligte

• Fonseca-de-Souza, André Luiz
• Freitas-Mesquita, Anita Leocadio
• Vieira, Lisvane Paes
• Majerowicz, David
• Daflon-Yunes, Nathalia
• Soares-de-Medeiros, Lia Carolina Almeida
• Miranda, Kildare
• Gondim, Katia Calp
• Meyer-Fernandes, José Roberto
• Rodrigues, Mauricio Martins

Titel

Identification and characterization of an ecto-pyrophosphatase activity in intact epimastigotes of Trypanosoma rangeli

Ist Teil von

• PloS one, 2014-09, Vol.9 (9), p.e106852-e106852

Ort / Verlag

United States: Public Library of Science

Erscheinungsjahr

2014

Quelle

MEDLINE

Beschreibungen/Notizen

• In this study, we performed the molecular and biochemical characterization of an ecto-enzyme present in Trypanosoma rangeli that is involved with the hydrolysis of extracellular inorganic pyrophosphate. PCR analysis identified a putative proton-pyrophosphatase (H(+)-PPase) in the epimastigote forms of T. rangeli. This protein was recognized with Western blot and flow cytometry analysis using an antibody against the H(+)-PPase of Arabidopsis thaliana. Immunofluorescence microscopy confirmed that this protein is located in the plasma membrane of T. rangeli. Biochemical assays revealed that the optimum pH for the ecto-PPase activity was 7.5, as previously demonstrated for other organisms. Sodium fluoride (NaF) and aminomethylenediphosphonate (AMDP) were able to inhibit approximately 75% and 90% of the ecto-PPase activity, respectively. This ecto-PPase activity was stimulated in a dose-dependent manner by MgCl2. In the presence of MgCl2, this activity was inhibited by millimolar concentrations of CaCl2. The ecto-PPase activity of T. rangeli decreased with increasing cell proliferation in vitro, thereby suggesting a role for this enzyme in the acquisition of inorganic phosphate (Pi). Moreover, this activity was modulated by the extracellular concentration of Pi and increased approximately two-fold when the cells were maintained in culture medium depleted of Pi. All of these results confirmed the occurrence of an ecto-PPase located in the plasma membrane of T. rangeli that possibly plays an important role in phosphate metabolism of this protozoan.