TY - JOUR T1 - Identification of structural and molecular features involved in the transport of 3'-deoxy-nucleoside analogs by human equilibrative nucleoside transporter 3 JF - Drug Metabolism and Disposition JO - Drug Metab Dispos DO - 10.1124/dmd.117.079400 SP - dmd.117.079400 AU - Rajgopal Govindarajan AU - Radhika Raj AU - MD Fazlur Rahman Y1 - 2018/01/01 UR - http://dmd.aspetjournals.org/content/early/2018/03/12/dmd.117.079400.abstract N2 - Combination antiretroviral drug treatments depend on 3'-deoxy-nucleoside analogs such as AZT (3'-azido-3'-deoxythymidine) and DDI (2'3'-dideoxyinosine). Despite being effective in inhibiting HIV viral replication, these drugs produce a range of toxicities, including myopathy, pancreatitis, neuropathy and lactic acidosis, that are generally considered as sequelae to mitochondrial damage. While the cell surface localized nucleoside transporters (e.g., human equilibrative nucleoside transporter 2 (hENT2), human concentrative nucleoside transporter 1 (hCNT1)) are known to increase the carrier mediated uptake of 3'-deoxy-nucleoside analogs into cells, another ubiquitously expressed intracellular nucleoside transporter, namely, hENT3, has been implicated in the mitochondrial transport of 3'-deoxy-nucleoside analogs. Using site-directed mutagenesis, generation of chimeric hENTs, and 3H-permeant flux measurements in mutant/chimeric-RNA injected Xenopus oocytes, here we identified the molecular determinants of hENT3 that dictate membrane translocation of 3'-deoxy-nucleoside analogs. Our findings demonstrated that whereas hENT1 had no significant transport activity towards 3'-deoxy-nucleoside analogs, hENT3 is capable of transporting 3'-deoxy-nucleoside analogs similar to hENT2. Transport analyses of hENT3-hENT1 chimeric constructs demonstrated that the N-terminal half of hENT3 is primarily responsible for the hENT3-3'-deoxy-nucleoside analogs interaction. In addition, mutagenic studies identified that 225D and 231L in the N-terminal half of hENT3 partially contribute to hENT3's ability to transport AZT and DDI. The identification of the transporter segment and amino acid residues that are important in hENT3 transport of 3'-deoxy-nucleoside analogs may present a possible mechanism for overcoming the adverse toxicities associated with 3'-deoxy-nucleoside analogs treatment and guide rational development of novel nucleoside analogs. ER -