High-resolution structures with bound Mn2+ and Cd2+ map the metal import pathway in an Nramp transporter

• Published in: eLife Vol. 12
• Main Authors: Ray, Shamayeeta, Berry, Samuel P, Wilson, Eric A, Zhang, Casey H, Shekhar, Mrinal, Singharoy, Abhishek, Gaudet, Rachelle
• Format: Journal Article
• Language: English
• Published: Cambridge eLife Sciences Publications Ltd 11-04-2023; eLife Sciences Publications, Ltd
• Subjects: Binding sites; Calorimetry; Deinococcus radiodurans; Homeostasis; isothermal titration calorimetry; Macrophages; Metabolism; Metal ions; Metals; Mutation; Nramp protein; Nramp transporters; Physiology; Protein transport; Spheres; Structural Biology and Molecular Biophysics; Titration; transition metal ion transport; x-ray crystallography
• ISSN: 2050-084X; 2050-084X
• DOI: 10.7554/eLife.84006

Transporters of the Nramp (Natural resistance-associated macrophage protein) family import divalent transition metal ions into cells of most organisms. By supporting metal homeostasis, Nramps prevent diseases and disorders related to metal insufficiency or overload. Previous studies revealed that Nramps take on a LeuT fold and identified the metal-binding site. We present high-resolution structures of
Deinococcus radiodurans
(Dra)Nramp in three stable conformations of the transport cycle revealing that global conformational changes are supported by distinct coordination geometries of its physiological substrate, Mn
2+
, across conformations, and by conserved networks of polar residues lining the inner and outer gates. In addition, a high-resolution Cd
2+
-bound structure highlights differences in how Cd
2+
and Mn
2+
are coordinated by DraNramp. Complementary metal binding studies using isothermal titration calorimetry with a series of mutated DraNramp proteins indicate that the thermodynamic landscape for binding and transporting physiological metals like Mn
2+
is different and more robust to perturbation than for transporting the toxic Cd
2+
metal. Overall, the affinity measurements and high-resolution structural information on metal substrate binding provide a foundation for understanding the substrate selectivity of essential metal ion transporters like Nramps.