Cloning of Leishmania nucleoside transporter genes by rescue of a transport-deficient mutant.

摘要: All parasitic protozoa studied to date are incapable of purine biosynthesis and must therefore salvage nucleobases or nucleosides from their hosts. This process is initiated by transporters on the parasite cell surface. We have used a mutant line (TUBA5) Leishmania donovani that deficient in adenosine/pyrimidine nucleoside transport activity (LdNT1) clone genes encoding these functional rescue. Two such genes, LdNT1.1 LdNT1.2, been sequenced shown encode deduced polypeptides with significant sequence identity human facilitative transporter hENT1. Hydrophobicity analysis LdNT1.2 proteins predicted 11 transmembrane domains. Transfection transport-deficient TUBA5 parasites vectors containing confers sensitivity cytotoxic adenosine analog tubercidin concurrently restores ability this take up [3H]adenosine [3H]uridine. Moreover, expression ORF Xenopus oocytes significantly increases [3H]adenosine, confirming single protein sufficient mediate transport. These results establish genetically biochemically both LdNT1 transporters.