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Structures in multiple conformations reveal distinct transition metal and proton pathways in an Nramp transporter.

<div class='caption-title'>Design and validation of complementary conformationally-locked crystallization constructs.</div>
<div class='caption-title'>Validation of conformationally-locked crystallization constructs.</div>
<div class='caption-title'>(A) Updated DraNramp Patch mutant structure in an inward-open apo state with a wide intracellular aqueous vestibule (red mesh) (Bozzi et al., 2016b). TMs 1, 5, 6, and 10 are colored gold, TMs 3, 4, 8, and 9 blue, and TMs 2, 7, and 11 gray. (B) The G45R structure revealed an apo inward-occluded conformation with no substantial extracellular or intracellular aqueous pathway to the metal-binding site, denoted by an * in structures where no metal is present. (C) The G223W structure revealed an outward-open conformation with the physiological substrate Mn2+ (magenta sphere) bound at the bottom of a substantial extracellular aqueous cavity (red mesh). All structures are viewed from within the membrane. (D) The complete Nramp transport cycle likely consists of at least six distinct conformational states. Including the structures of S. capitis (Sca) (Ehrnstorfer et al., 2014) and E. coleocola (Eco) (Ehrnstorfer et al., 2017) homologs, we now have structures of five of these conformations. (E) Pairwise RMSD values for the superposition of the 355 Cα atoms present in all three DraNramp structures, and ScaNramp and EcoNramp. G223W superimposes best with the outward-open EcoNramp, validating our mutagenesis strategy to obtain an outward-open conformation. G45R superimposes better with the inward-open Patch mutant than outward-open G223W, suggesting it represents an inward-occluded conformation. (F) Superposition of Patch mutant (gold, gray, and blue) and G45R (cyan) and (G) RMSD calculated at each Cα. These structures superimpose well except for TM1a. TM1a’s significant displacement from the inward-occluded state (G45R) to the inward-open state (Patch) is highlighted with a red arrow. (H) Superposition of G223W (gold, gray, and blue) and G45R (cyan) and (I) RMSD calculated at each Cα. The most significant rearrangements involve TMs 1b, 4, 5, 6a, and 10. These significant displacements from the inward-occluded state (G45R) to the outward-open state (G223W) are highlighted with red arrows. In panels G and I, black lines indicate average RMSD for each TM, with TMs 1, 6, and 10 divided into two halves. In panels F and H, the central view is rotated 45° along the vertical axis from the view in panels A-C, while the left and right views are 90° rotations of that central view. See also Figure 2—figure supplements 1 and ​and22.</div>
Bozzi AT, Zimanyi CM, Nicoludis JM, Lee BK, Zhang CH, Gaudet R
OpenAccess logo Elife 8 (2019)
Related entries: 6d91, 6d9w

Cited: 30 times
EuropePMC logo PMID: 30714568

Abstract

Nramp family transporters-expressed in organisms from bacteria to humans-enable uptake of essential divalent transition metals via an alternating-access mechanism that also involves proton transport. We present high-resolution structures of Deinococcus radiodurans (Dra)Nramp in multiple conformations to provide a thorough description of the Nramp transport cycle by identifying the key intramolecular rearrangements and changes to the metal coordination sphere. Strikingly, while metal transport requires cycling from outward- to inward-open states, efficient proton transport still occurs in outward-locked (but not inward-locked) DraNramp. We propose a model in which metal and proton enter the transporter via the same external pathway to the binding site, but follow separate routes to the cytoplasm, which could facilitate the co-transport of two cationic species. Our results illustrate the flexibility of the LeuT fold to support a broad range of substrate transport and conformational change mechanisms.

Reviews - 6c3i mentioned but not cited (3)

  1. Molecular Mechanism of Nramp-Family Transition Metal Transport. Bozzi AT, Gaudet R. J Mol Biol 433 166991 (2021)
  2. Ins and Outs: Recent Advancements in Membrane Protein-Mediated Prokaryotic Ferrous Iron Transport. Brown JB, Lee MA, Smith AT. Biochemistry 60 3277-3291 (2021)
  3. Structural features and functional implications of proteins enabling the robustness of Deinococcus radiodurans. Chen Z, Tang Y, Hua Y, Zhao Y. Comput Struct Biotechnol J 18 2810-2817 (2020)

Articles - 6c3i mentioned but not cited (5)

  1. Structures in multiple conformations reveal distinct transition metal and proton pathways in an Nramp transporter. Bozzi AT, Zimanyi CM, Nicoludis JM, Lee BK, Zhang CH, Gaudet R. Elife 8 e41124 (2019)
  2. Transmembrane helix 6b links proton and metal release pathways and drives conformational change in an Nramp-family transition metal transporter. Bozzi AT, McCabe AL, Barnett BC, Gaudet R. J Biol Chem 295 1212-1224 (2020)
  3. Structural and functional properties of a magnesium transporter of the SLC11/NRAMP family. Ramanadane K, Straub MS, Dutzler R, Manatschal C. Elife 11 e74589 (2022)
  4. High-resolution structures with bound Mn2+ and Cd2+ map the metal import pathway in an Nramp transporter. Ray S, Berry SP, Wilson EA, Zhang CH, Shekhar M, Singharoy A, Gaudet R. Elife 12 e84006 (2023)
  5. Structural and functional properties of a plant NRAMP-related aluminum transporter. Ramanadane K, Liziczai M, Markovic D, Straub MS, Rosalen GT, Udovcic A, Dutzler R, Manatschal C. Elife 12 e85641 (2023)


Reviews citing this publication (3)

  1. General principles of secondary active transporter function. Beckstein O, Naughton F. Biophys Rev (Melville) 3 011307 (2022)
  2. Deciphering the functional roles of transporter proteins in subcellular metal transportation of plants. Chen X, Zhao Y, Zhong Y, Chen J, Qi X. Planta 258 17 (2023)
  3. Structures and coordination chemistry of transporters involved in manganese and iron homeostasis. Ray S, Gaudet R. Biochem Soc Trans 51 897-923 (2023)

Articles citing this publication (19)

  1. Unique structural features in an Nramp metal transporter impart substrate-specific proton cotransport and a kinetic bias to favor import. Bozzi AT, Bane LB, Zimanyi CM, Gaudet R. J Gen Physiol 151 1413-1429 (2019)
  2. Heavy Metal Transporters-Associated Proteins in S. tuberosum: Genome-Wide Identification, Comprehensive Gene Feature, Evolution and Expression Analysis. He G, Qin L, Tian W, Meng L, He T, Zhao D. Genes (Basel) 11 E1269 (2020)
  3. Mechanistic basis of the inhibition of SLC11/NRAMP-mediated metal ion transport by bis-isothiourea substituted compounds. Manatschal C, Pujol-Giménez J, Poirier M, Reymond JL, Hediger MA, Dutzler R. Elife 8 e51913 (2019)
  4. Manganese impairs the QoxABCD terminal oxidase leading to respiration-associated toxicity. Sachla AJ, Luo Y, Helmann JD. Mol Microbiol 116 729-742 (2021)
  5. The two-domain elevator-type mechanism of zinc-transporting ZIP proteins. Wiuf A, Steffen JH, Becares ER, Grønberg C, Mahato DR, Rasmussen SGF, Andersson M, Croll T, Gotfryd K, Gourdon P. Sci Adv 8 eabn4331 (2022)
  6. A solid-supported membrane electrophysiology assay for efficient characterization of ion-coupled transport. Thomas NE, Feng W, Henzler-Wildman KA. J Biol Chem 297 101220 (2021)
  7. Integrated AlphaFold2 and DEER investigation of the conformational dynamics of a pH-dependent APC antiporter. Del Alamo D, DeSousa L, Nair RM, Rahman S, Meiler J, Mchaourab HS. Proc Natl Acad Sci U S A 119 e2206129119 (2022)
  8. Unconventional transport of metal ions and protons by Nramps. Rudnick G. J Gen Physiol 151 1339-1342 (2019)
  9. Activating alternative transport modes in a multidrug resistance efflux pump to confer chemical susceptibility. Spreacker PJ, Thomas NE, Beeninga WF, Brousseau M, Porter CJ, Hibbs KM, Henzler-Wildman KA. Nat Commun 13 7655 (2022)
  10. Divalent metal transporter-related protein restricts animals to marine habitats. Sassa M, Takagi T, Kinjo A, Yoshioka Y, Zayasu Y, Shinzato C, Kanda S, Murakami-Sugihara N, Shirai K, Inoue K. Commun Biol 4 463 (2021)
  11. Electrophysiology Measurements of Metal Transport by MntH2 from Enterococcus faecalis. Gantner M, Laftsoglou T, Rong H, Postis VLG, Jeuken LJC. Membranes (Basel) 10 E255 (2020)
  12. Nramp: Deprive and conquer? Cellier MFM. Front Cell Dev Biol 10 988866 (2022)
  13. PcNRAMP1 Enhances Cadmium Uptake and Accumulation in Populus × canescens. Yu W, Deng S, Chen X, Cheng Y, Li Z, Wu J, Zhu D, Zhou J, Cao Y, Fayyaz P, Shi W, Luo Z. Int J Mol Sci 23 7593 (2022)
  14. Structural and Functional Characterization of the Holliday Junction Resolvase RuvC from Deinococcus radiodurans. Qin C, Han W, Xu Y, Zhao Y, Xu H, Tian B, Wang L, Hua Y. Microorganisms 10 1160 (2022)
  15. Structures of ferroportin in complex with its specific inhibitor vamifeport. Lehmann EF, Liziczai M, Drożdżyk K, Altermatt P, Langini C, Manolova V, Sundstrom H, Dürrenberger F, Dutzler R, Manatschal C. Elife 12 e83053 (2023)
  16. The Conserved Serine Transporter SdaC Moonlights To Enable Self Recognition. Chittor A, Gibbs KA. J Bacteriol 204 e0034721 (2022)
  17. Charge neutralization of the active site glutamates does not limit substrate binding and transport by small multidrug resistance transporter EmrE. Spreacker PJ, Brousseau M, Hisao GS, Soltani M, Davis JH, Henzler-Wildman KA. J Biol Chem 299 102805 (2023)
  18. Slc11 Synapomorphy: A Conserved 3D Framework Articulating Carrier Conformation Switch. Cellier MFM. Int J Mol Sci 24 15076 (2023)
  19. Structural basis of ion uptake in copper-transporting P1B-type ATPases. Salustros N, Grønberg C, Abeyrathna NS, Lyu P, Orädd F, Wang K, Andersson M, Meloni G, Gourdon P. Nat Commun 13 5121 (2022)


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