IDENTIFICATION AND CHARACTERIZATION OF A FUNCTIONAL PEROXIDOXIN FROM LEISHMANIA-MAJOR

Leishmania spp, encounter damaging oxygen metabolites from endogenous metabolic processes as well as from exogenous sources, such as inside the gut of the sandfly vector and within host macrophages. The recentl y described peroxidoxin protein family form part of a novel pathway fo r metabolising hydrogen peroxide that, in trypanosomatids, links perox ide reduction to NADPH oxidation via trypanothione. Here we report the cloning and characterisation of the Leishmania major peroxidoxin gene , tryparedoxin peroxidase (TryP). TryP is a multi-copy gene arranged i n a complex tandem array located on the size polymorphic homologues of chromosome 15. Northern analysis showed that TryP expresses a single 1.6 kb mRNA throughout promastigote development. TryP encodes a 22-kDa protein with two conserved cysteine-containing domains that defines i t as a 2-Cys peroxidoxin. Purified recombinant TryP protein catabolise d hydrogen peroxide in the presence of the tryparedoxin homologue from Crithidia fasciculata (Cf-TryX), trypanothione, trypanothione reducta se and NADPH. The demonstration that L. major utilises a three-protein peroxidase system confirms that this is a mechanism of protection aga inst oxidative damage in this parasite. (C) 1998 Elsevier Science B.V. All rights reserved.