br The glyoxalase system has

The glyoxalase system has not yet been studied in E. histolytica or G. lamblia. Searches in the G. lamblia genome (GiardiaDB.org) revealed two genes encoding hydroxyacylglutathione hydrolases (Morrison et al., 2007), but no glyoxalase I gene was found. The genome of E. histolytica does not have a GLO1, and there are no annotated GLO2-encoding genes (AmoebaDB.org). Instead, this parasite has nine genes coding for proteins that belong to the metallo-β-lactamase family (AmoebaDB.org). These parasites have an unusual thiol metabolism, lacking glutathione and glutathione-dependent enzymes, as in glutathione reductase and glutathione peroxidase (reviewed in Thompson et al., 1993). In these parasites, cysteine is the main low-molecular-mass thiol (Krauth-Siegel and Leroux, 2012). In the absence of GLO1 and glutathione, the elimination of methylglyoxal through the glyoxalase system is highly unlikely to occur in these parasites. Additionally, G. lamblia and E. histolytica have two and three genes, respectively, that encode for aldose reductase (EC 1.1.1.21) and may be responsible for the NADPH-dependent methylglyoxal detoxification.
The glyoxalase pathway was first identified in Leishmania braziliensis as a system that converts methylglyoxal into d-lactate using reduced glutathione (Darling and Blum, 1988). Several years later, the characterization of glyoxalase I and II in different trypanosomatids revealed that trypanothione, a bis(glutathione)–spermidine conjugate, was the physiological substrate for these GSK2292767 (Irsch and Krauth-Siegel, 2004, Sousa Silva et al., 2005, Vickers et al., 2004). Glyoxalase I and II are found in the genomes of all Leishmania species (TriTrypDB.org). Also in T. cruzi there is one gene encoding for glyoxalase I (UniProt Q4D7B4; Greig et al., 2006) and one encoding a putative GLO2 (UniProt Q4E0K0). However, the absence of a GLO1 gene in T. brucei (Wendler et al., 2009) and the existence of two putative GLO2 genes, only one encoding a functional glyoxalase II enzyme, is intriguing (UniProt Q6KF36; (Irsch and Krauth-Siegel, 2004, Wendler et al., 2009). Lacking glyoxalase I, it is assumed that methylglyoxal elimination in T. brucei mainly occurs through the methylglyoxal reductase pathway, resulting in l-lactate (Greig et al., 2009). To date, there are only two trypanosomatid glyoxalase I-solved structures: from L. major (UniProt Q68RJ8; Ariza et al., 2006) and L. infantum (UniProt A4IBI9; Barata et al., 2010). They are structurally very similar, and both are dimeric proteins (three dimers are present in the asymmetric unit of the orthorhombic crystal form, unpublished data for LiGLO1). Binding the substrate through its glutathione moiety (Ariza et al., 2006), this enzyme can also react with the glutathione-derived hemithioacetal, although with less affinity (Sousa Silva et al., 2005, Vickers et al., 2004). In L. infantum GLO1 structure (Barata et al., 2010), both trypanothione and glutathione-dependent substrates can be accommodated, but the active site allows a better fitting for the first (our work, unpublished data). Like all known glyoxalases I, trypanosomatid enzymes contain a metal center. Nickel, found in prokaryotic glyoxalase I enzymes (Sukdeo et al., 2004), is present in the GLO1 of T. cruzi (Greig et al., 2006) and L. major (Ariza et al., 2006, Vickers et al., 2004), and a prokaryotic origin was assumed for L. major glyoxalase I based on metal binding specificity and the high degree of sequence similarity with E. coli GLO1 (Vickers et al., 2004). This is an exception, since zinc is generally found in eukaryotic enzymes, including human (Cameron et al., 1997), yeast (Aronsson et al., 1978), and P. falciparum (Iozef et al., 2003). Interestingly, glyoxalase I from L. infantum contains zinc at the metal binding site (our work, unpublished results), as do the eukaryotic GLO1 enzymes, but the relevance of this metal specificity change in two closely related species, L. infantum and L. major, is still questionable since the enzymes are very similar, both catalytically and structurally.