Trypanothione-dependent peroxide metabolism in Trypanosoma cruzi different stages

Abstract

Different stages of Trypanosoma cruzi are able to metabolize low concentrations of H 2O 2. Trypomastigotes showed a higher initial rate per mg protein than amastigotes or epimastigotes derived from them. Amastigotes could metabolize H 2O 2 at a lower rate than the other developmental stages of T. cruzi. A peroxide-metabolizing activity was detected in extracts of T. cruzi epimastigotes. This ‘NADPH peroxidase’ activity was lost upon dialysis of the extracts and was probably due to a non-enzymatic reaction(s) with endogenous dihydrotrypanothione (T(SH) 2) and/or other thiols, thus explaining the inhibition of H 2O 2 metabolism in intact cells by thiol inhibitors. An amount of non-protein thiols equivalent to an intracellular concentration of 2.0–3.0 mM was found in epimastigotes, which is sufficient to account for the rate of NADPH oxidation observed in the presence of high concentration of peroxides (> 100 μM). Addition of T(SH) 2 increased this rate, implying that this thiol could be used as a substrate in that reaction. In addition, this activity was hardly detectable in the extracts in the presence of low concentration of peroxides (<20 μM), indicating a high K m, which would be incompatible with a true peroxidase activity. Taking into account the high intracellular concentration of thiols measured, this activity probably accounted for the rates of H 2O 2 metabolism detected in intact T. cruzi. These results also confirm that T. cruzi is an organism with limited ability to detoxify H 2O 2.