This article provides an overview about the recent advances in the dissection of the peroxide metabolism of Trypanosomatidae. This family of protozoan organisms comprises the medically relevant parasites Trypanosoma brucei, Trypanosoma cruzi, and Leishmania spp. Over the past 10 years, three major families of peroxidases have been identified in these organisms: (a) 2-cysteine peroxiredoxins, (b) nonselenium glutathione peroxidases, and (c) ascorbate peroxidases. In trypanosomatids, these enzymes display the unique feature of using reducing equivalents derived from trypanothione, a dithiol found exclusively in these protozoa. The electron transfer between trypanothione and the peroxidases is mediated by a redox shuttle, which can either be tryparedoxin, ascorbate, or even glutathione. The preference for the intermediate molecule differs among each peroxidase and so does the specificity for the peroxide substrate. These observations, added to the fact that these peroxidases are distributed throughout different subcellular compartments, point to the existence of an elaborate peroxide metabolism in trypanosomatids. With the completion of the trypanosomatids genome, other molecules displaying peroxidase activity might be added to this list in the future.

Significance: Peroxiredoxins (Prxs) are highly abundant proteins, which serve two seemingly mutually exclusive roles as peroxidases and molecular chaperones. Little is known about the precise mechanism of Prxs’ activation as chaperone and the physiological significance of this second function. Here we demonstrate that inLeishmania infantum, reduced Prx provides a crucial, stress-specific chaperone reservoir, which is activated rapidly upon exposure to unfolding stress conditions. Once activated, Prx protects a wide range of different clients against protein unfolding. Clients are bound in the center of the decameric ring, providing experimental evidence for previous claims that Prxs serve as likely ancestors of chaperonins. Interference with client binding impairsLeishmaniainfectivity, providing compelling evidence for the in vivo importance of Prx’s chaperone function.

Two-cysteine peroxiredoxins are ubiquitous peroxidases that play various functions in cells. In Leishmania and related trypanosomatids, which lack catalase and selenium-glutathione peroxidases, the discovery of this family of enzymes provided the molecular basis for peroxide removal in these organisms. In this report the functional relevance of one of such enzymes, the mitochondrial 2-Cys peroxiredoxin (mTXNPx), was investigated along the Leishmania infantum life cycle. mTXNPx null mutants (mtxnpx⁻) produced by a gene replacement strategy, while indistinguishable from wild type promastigotes, were found unable to thrive in a murine model of infection. Unexpectedly, however, the avirulent phenotype of mtxnpx⁻ was not due to lack of the peroxidase activity of mTXNPx as these behaved like controls when exposed to oxidants added exogenously or generated by macrophages during phagocytosis ex vivo. In line with this, mtxnpx⁻ were also avirulent when inoculated into murine hosts unable to mount an effective oxidative phagocyte response (B6.p47^phox−/− and B6.RAG2^−/− IFN-γ^−/− mice). Definitive conclusion that the peroxidase activity of mTXNPx is not required for parasite survival in mice was obtained by showing that a peroxidase-inactive version of this protein was competent in rescuing the non-infective phenotype of mtxnpx⁻. A novel function is thus proposed for mTXNPx, that of a molecular chaperone, which may explain the impaired infectivity of the null mutants. This premise is based on the observation that the enzyme is able to suppress the thermal aggregation of citrate synthase in vitro. Also, mtxnpx⁻ were more sensitive than controls to a temperature shift from 25°C to 37°C, a phenotype reminiscent of organisms lacking specific chaperone genes. Collectively, the findings reported here change the paradigm which regards all trypanosomatid 2-Cys peroxiredoxins as peroxide-eliminating devices. Moreover, they demonstrate, for the first time, that these 2-Cys peroxiredoxins can be determinant for pathogenicity independently of their peroxidase activity. Leishmania are insect-borne protozoan parasites that cause leishmaniasis, a spectrum of diseases with relevance in men and dogs and for which there are no adequate means of control. In order to successfully infect their mammalian hosts, Leishmania requires the expression of a set of key proteins. Here, we identify one mitochondrial protein of the parasite as one of such molecules. Interestingly, however, we also discovered that the reason why this enzyme is essential is not due to its well characterized peroxidase activity, which defines it as a member of the peroxiredoxin family. Instead, the importance of this protein during infection may derive from its ability to function as a chaperone, as implied here. In short, by proposing a novel, peroxidase-unrelated role for the Leishmania mitochondrial peroxiredoxin during mammals' infection, this work opens new perspectives regarding the physiologic function of this enzyme.