Structural and functional properties of a magnesium transporter of the SLC11/NRAMP family

Abstract

Members of the ubiquitous SLC11/NRAMP family catalyze the uptake of divalent transition metal ions into cells. They have evolved to efficiently select these trace elements from a large pool of Ca2+ and Mg2+, which are both orders of magnitude more abundant, and to concentrate them in the cytoplasm aided by the cotransport of H+ serving as energy source. In the present study, we have characterized a member of a distant clade of the family found in prokaryotes, termed NRMTs, that were proposed to function as transporters of Mg2+. The protein transports Mg2+ and Mn2+ but not Ca2+ by a mechanism that is not coupled to H+. Structures determined by cryo-EM and X-ray crystallography revealed a generally similar protein architecture compared to classical NRAMPs, with a restructured ion binding site whose increased volume provides suitable interactions with ions that likely have retained much of their hydration shell.

Nramp: Deprive and conquer?

“…The outgroup exhibits only 1 conserved charge part of Slc11 H + -network, in TMS3 (ancestral Glu), which assumes distinct orientations in NRMT and MntH (Supplementary Figure S8) and rearranges upon Mn 2+ binding by MntH ( Figure 3A ). Prior to MB emergence, 5 co-evolved sites in TMS1&6 (Asp, Asn and Ala, Met, His, respectively) enabled both counter-selecting Mg 2+ , most likely due to steric constraint preventing the binding a hydrated Mg 2+ ion, and allowing instead Mn 2+ import coupled to H + uptake ( Chaloupka et al, 2005 ; Ehrnstorfer et al, 2014 ; Makui et al, 2000 ; Ramanadane et al, 2022 ). Toward MA, 4 novel charges were selected in TMS3&9 (Glu, Asp and Arg, Arg, respectively) to create a divergent H + translocation pathway linked to TMS3 ancestral charge ( Bozzi et al, 2019a ).…”

“…Stephane Rety, INSERM U1210 Laboratoire de Biologie et Modelisation de la Cellule (LBMC), France (Cellier, 2012b;Ehrnstorfer et al, 2014;Ponzoni et al, 2018;Ramanadane et al, 2022;Shin et al, 2014;Vastermark et al, 2014;Zallot et al, 2019). Slc11-specific residues in transmembrane segments (TMS) 1&6 are key to H +dependent import of divalent transition metals (e.g., Mn(II), Fe(II), Co(II)), protons co-transport providing the motive force necessary to import Mn(II) against its electro-chemical gradient.…”

“…Slc11 3D architecture fits the “LeuT-fold,” a structural paradigm for the Amino acid-Polyamine-organoCation (APC) superfamily of chemiosmotic transporters or carriers (Pfam Clan CL0062). This Pfam Clan includes the family PF01566, which comprises both the Slc11 family and its phylogenetic outgroup that shows less than 30% sequence identity as well as distinct catalytic specificities (e.g., Nramp-related Mg 2+ transporters, NRMT) ( Cellier, 2012b ; Ehrnstorfer et al, 2014 ; Ponzoni et al, 2018 ; Ramanadane et al, 2022 ; Shin et al, 2014 ; Vastermark et al, 2014 ; Zallot et al, 2019 ). Slc11-specific residues in transmembrane segments (TMS) 1&6 are key to H + -dependent import of divalent transition metals (e.g., Mn(II), Fe(II), Co(II)), protons co-transport providing the motive force necessary to import Mn(II) against its electro-chemical gradient.…”

“…Slc11-specific residues in transmembrane segments (TMS) 1&6 are key to H + -dependent import of divalent transition metals (e.g., Mn(II), Fe(II), Co(II)), protons co-transport providing the motive force necessary to import Mn(II) against its electro-chemical gradient. In contrast, the corresponding NRMT-specific residues facilitate instead H + -independent uptake of Mg(II), thereby indicating type II coevolutionary rate-shifted sites that defined Slc11 specificity ( Courville et al, 2008 ; Ehrnstorfer et al, 2014 ; Ramanadane et al, 2022 ).…”

High-resolution structures map the metal import pathway in an Nramp transporter

“…The metal-binding methionine is essential to select against alkaline earth metals 19 . This finding is corroborated by the fact that a bacterial Nramp homolog which lacks a metal-binding methionine, NRMT, can transport Mg 2+ (ref 20 ). However, little is known about whether from their promiscuous spectrum of transition metal substrates, the canonical Nramps can mechanistically distinguish between their physiological substrates (Fe 2+ and Mn 2+ ) from non-essential ones like Cd 2+ .…”