Biochemical characterization of a trypanosome enzyme with glutathione-dependent peroxidase activity

In most eukaryotes, glutathione-dependent peroxidases play a key role in th e metabolism of peroxides. Numerous studies have reported that trypanosomat ids lack this activity. Here we show that this is not the case, at least fo r the American trypanosome Trypanosoma cruzi. We have isolated a single-cop y gene from T. cruzi with the potential to encode an 18 kDa enzyme, the seq uence of which has highest similarity with glutathione peroxidases from pla nts. A recombinant form of the protein was purified following expression in Escherichia coli, The enzyme was shown to have peroxidase activity in the presence of glutathione/glutathione reductase but not in the presence of tr ypanothione/trypanothione reductase. It could metabolize a wide range of hy droperoxides (linoleic acid hydroperoxide and phosphatidyl-choline hydroper oxide > cumenehydroperoxide > t-butylhydroperoxide), but no activity toward s hydrogen peroxide was detected. Enzyme activity could be saturated by glu tathione when both fatty acid and short-chain organic hydroperoxides were u sed as substrate. For linoleic acid hydroperoxide, the rate-limiting step o f this reaction is the reduction of the peroxidase by glutathione. With low er-affinity substrates such as t-butyl hydroperoxide, the rate-limiting ste p is the reduction of the oxidant. The data presented here identify a new a rm of the T, cruzi oxidative defence system.