Leishmania amazonensis: Effects of heat shock on ecto-ATPase activity

In this work we demonstrated that promastigotes of Leishmania amazonensis exhibit an Mg-dependent ecto-ATPase activity, which is stimulated by heat shock. The Mg-dependent ATPase activity of cells grown at 22 and 28 °C was 41.0 ± 5.2 nmol Pi/h × 107 cells and 184.2 ± 21.0 nmol Pi/h × 107 cells, respectively. When both promastigotes were pre-incubated at 37 °C for 2 h, the ATPase activity of cells grown at 22 °C was increased to 136.4 ± 10.6 nmol Pi/h × 107 whereas that the ATPase activity of cells grown at 28 °C was not modified by the heat shock (189.8 ± 10.3 nmol Pi/h × 107 cells). It was observed that Km of the enzyme from cells grown at 22 °C (Km = 980.2 ± 88.6 μM) was the same to the enzyme from cells grown at 28 °C (Km = 901.4 ± 91.9 μM). In addition, DIDS (4,4′-diisothiocyanatostilbene 2,2′-disulfonic acid) and suramin, two inhibitors of ecto-ATPases, also inhibited similarly the ATPase activities from promastigotes grown at 22 and 28 °C. We also observed that cells grown at 22 °C exhibit the same ecto-phosphatase and ecto 3′- and 5′-nucleotidase activities than cells grown at 28 °C. Interestingly, cycloheximide, an inhibitor of protein synthesis, suppressed the heat-shock effect on ecto-ATPase activity of cells grown at 22 °C were exposed at 37 °C for 2 h. A comparison between the stimulation of the Mg-dependent ecto-ATPase activity of virulent and avirulent promastigotes by the heat shock showed that avirulent promastigotes had a higher stimulation than virulent promastigotes after heat stress.