Functional properties of a prototypical member of the NRAMP superfamily of metal‐ion transporters

Human NRAMP1 (SLC11A1) and DMT1 (SLC11A2) metal‐ion transporters play nonredundant roles in host defense against infection and in iron homeostasis, respectively. Both are archetypical members of a superfamily extending to plants, yeasts and bacteria. The existence of prototypical and archetypical NRAMP homologs in simple eukaryotic organisms provides the opportunity to explore functional divergence and evolution in this family. We are characterizing a prototype NRAMP homolog ( Pp pNRAMP) from an ancestral green moss, Physcomitrella patens . Disk‐diffusion assays upon expression of Pp pNRAMP in E. coli revealed increased cytotoxicity in the presence of Co and Cd, but not Fe, consistent with bacterial 109 Cd 2+ uptake. In Xenopus oocytes expressing Pp pNRAMP, we observed at alkaline pH o large constitutive inward currents that were reversed in the presence of divalent metal ions, ranked Zn 2+ , Cd 2+ ( K 0.5 ≈ 0.3 mM), Pb 2+ > Ni 2+ , Co 2+ , Mn 2+ , Sn 2+ >> Sr 2+ , Fe 2+ . Radiotracer assays revealed limited capacity for 109 Cd 2+ transport. Whereas human DMT1‐mediated Cd 2+ ‐evoked currents were temperature‐dependent ( E a = 14.4 ± 0.6 kcal.mol 1 ), those for Pp pNRAMP were less so ( E a = 2.8 ± 2.0 kcal.mol 1 ), suggestive of channel activity; however, a reversal‐potential approach has not yet identified the predominant ion conductance. Pp pNRAMP may serve as a paradigm for the NRAMP prototype subfamily. Unlike archetypical NRAMP proteins, Pp pNRAMP is not reactive with iron. Examining functional divergence between proto‐ and archetypical NRAMP homologs will help us identify key residues in human NRAMP1 and DMT1 proteins.