title: A Leishmania major nucleobase transporter responsible for allopurinol uptake is a functional homolog of the Trypanosoma brucei H2 transporter.
abstract: Nucleobase transporters play an important role in the physiology of protozoan parasites, because these organisms are purine auxotrophs and rely entirely on salvage of these vital compounds. Purine transporters have also been shown to mediate the uptake of important antiparasitic drugs. In the current study, we investigated the uptake of [(3)H]adenine, [(3)H]hypoxanthine, and [(3)H]allopurinol, an antileishmanial hypoxanthine analog, by Leishmania major. These compounds were all taken up by a single high-affinity transporter, LmNBT1, with K(m) values of 4.6 +/- 0.9, 0.71 +/- 0.07, and 54 +/- 3 microM, respectively. Guanine and xanthine fully inhibited [(3)H]adenine transport, with K(i) values of 2.8 +/- 0.7 and 23 +/- 8 microM. Using purine analogs, an inhibitor profile for LmNBT1 was obtained, which allowed the construction of a quantitative model for the interactions between the transporter binding site and the permeant. The model predicts that hypoxanthine was bound through hydrogen bonds to N(1)H, N3, N7, and N(9)H of the purine ring, with a total Gibbs free energy of -39.5 kJ/mol. The interactions with adenine were similar, except for a weak hydrogen bond to N1 (unprotonated in adenine). The predicted mode of substrate binding for LmNBT1 was almost identical to that for the Trypanosoma brucei H2 (TbH2) transporter. It is proposed that the architecture of their respective binding sites is very similar and that LmNBT1 can be named a functional homolog of TbH2.
authors.affiliation: Institute of Biomedical and Life Sciences, Division of Infection and Immunity, Joseph Black Building, University of Glasgow, Glasgow, United Kingdom.
authors.forename: Mohammed I
authors.initials: MI
authors.lastname: Al-Salabi
authors.forename: Lynsey J M
authors.initials: LJ
authors.lastname: Wallace
authors.forename: Harry P
authors.initials: HP
authors.lastname: De Koning
mesh_terms: D000225:Adenine / Q000378:metabolism; D000493:Allopurinol / Q000493:pharmacokinetics*; D000818:Animals; D001692:Biological Transport; D019271:Hypoxanthine / Q000378:metabolism; D018320:Leishmania major / Q000378:metabolism*; D033702:Nucleobase Transport Proteins / Q000737:chemistry / Q000378:metabolism*; D013329:Structure-Activity Relationship; D014316:Tritium; D014346:Trypanosoma brucei brucei / Q000737:chemistry / Q000378:metabolism
chemical_list: D033702:Nucleobase Transport Proteins; D014316:Tritium; D019271:Hypoxanthine; D000493:Allopurinol; D000225:Adenine