A new report in Proceedings of the National Academies of Sciences, USA, describes a prokaryotic-like glyoxalase pathway in the eukaryotic parasite Leishmania major, a finding that could provide a new target for rational drug design.

The glyoxalase pathway — which comprises the enzymes glyoxalase I and II (GLO1 and GLO2) — is required for the detoxification of endogenous toxins such as methylglyoxal. Methylglyoxal spontaneously forms a hemithioacetal adduct with glutathione, which is then converted by GLO1 and GLO2 into D-lactate and free glutathione.

Alan Fairlamb and co-workers identified a putative GLO1 gene in the recently completed L. major genome. Sequence comparisons of this gene revealed 47% identity to Escherichia coli, 51% identity to Synechococcus sp. WH 8102, but only 33% identity to human GLO1 sequences. In addition, phylogenetic analyses placed L. major GLO1 within the proteobacterial and cyanobacterial lineages, underlying the similarity to prokaryotic GLO1 enzymes.

GLO1 requires the presence of a metal co-factor for activity, and prokaryotes and eukaryotes have different requirements — human enzymes require zinc and E. coli enzymes require nickel. Analysis of the gene sequences showed that the residues shown to be important in E. coli metal binding are conserved in the L. major GLO1 sequence, indicating that L. major GLO1 requires nickel rather than zinc. To confirm this, the authors expressed and purified L. major GLO1 and stripped it of all metal co-factors. The resulting apoenzyme was reactivated with nickel, but zinc had no effect.

However, this proposed L. major glyoxalase pathway is also distinct to those of prokaryotes such as E. coli. L. major is known to have an unusual thiol metabolism — it produces trypanothione rather than glutathione. The authors looked at the kinetic properties of L. major GLO1 and found that, with trypanothione, it was as efficient as the human enzyme was with glutathione; however, there was a 200-fold difference between its activity with trypanothione and glutathione.

Taken together, these data provide evidence for a new category of trypanosomal GLO1 enzymes. New antitrypanosomal agents are needed, particularly in the developing world, and the differences between L. major and human glyoxalase systems indicate that it could be a much-needed target for rational drug design.