Purine nucleobase transport in amastigotes of Leishmania mexicana: involvement in allopurinol uptake.

Abstract

Nucleobase and nucleoside transporters play central roles in the biochemistry of parasitic protozoa, as they lack the ability to synthesize purines de novo and are absolutely reliant upon purine salvage from their hosts. Furthermore, such transporters are potentially critical to the pharmacology of these important human pathogens, because they mediate the uptake of purine analogues, as well as some nonpurine drugs, that can be selectively cytotoxic to the parasites. We here report the first identification and characterization of a purine nucleobase transporter in Leishmania amastigotes. Uptake of [3H]hypoxanthine by Leishmania mexicana amastigotes was mediated by a single high-affinity transporter, LmexNBT1, with a Km of 1.6 +/- 0.4 microM and high affinity for adenine, guanine, and xanthine but low affinity for nucleosides and pyrimidine nucleobases. Allopurinol, an antileishmanial hypoxanthine analogue, was apparently taken up by the same transporter. Using [3H]allopurinol, a Km value of 33.6 +/- 6.0 microM was obtained. All evidence was compatible with a model of a single purine nucleobase transporter being expressed in amastigotes. Using various purine nucleobase analogues, a model for the interactions between hypoxanthine and the transporter's permeant binding site was constructed. The binding interactions were compared with those of the LmajNBT1 transporter in Leishmania major promastigotes and found to be very similar.