Structural basis for alternating access of a eukaryotic calcium/proton exchanger

Waight, Andrew B.; Pedersen, Bjørn Panyella; Schlessinger, Avner; Bonomi, Massimiliano; Chau, Bryant; Roe-Žurž, Zygy; Risenmay, Aaron J.; Šali, Andrej; Stroud, Robert M.

doi:10.1038/nature12233

Cited by 94 publications

(120 citation statements)

References 54 publications

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“…We conclude, therefore, that if one of these configurations were indeed present in the protein crystal, alongside the three-Na + state described above, it would be minimally populated and thus would not be reflected in the electron density detected for the protein (although it may be sufficiently populated to yield a weak anomalous scattering signal). However, it is also possible that Ca 2+ binds in a configuration not considered here, for example, involving multiple water molecules mediating ion-protein contacts, as observed in a recent X-ray structure of a Ca 2+ /H + exchanger in the inward-facing conformation (29). The experimental electron density for NCX_Mj, however, reveals no indication of such a water network, so this configuration too would be marginally populated in the crystal.…”

Section: Simulations Identify Three Ion Configurations Similarly Consmentioning

confidence: 77%

Sodium recognition by the Na ⁺ /Ca ²⁺ exchanger in the outward-facing conformation

Marinelli

Almagor

Hiller

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

124

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Na + /Ca 2+ exchangers (NCXs) are ubiquitous membrane transporters with a key role in Ca 2+ homeostasis and signaling. NCXs mediate the bidirectional translocation of either Na + or Ca 2+ , and thus can catalyze uphill Ca 2+ transport driven by a Na + gradient, or vice versa. In a major breakthrough, a prokaryotic NCX homolog (NCX_Mj) was recently isolated and its crystal structure determined at atomic resolution. The structure revealed an intriguing architecture consisting of two inverted-topology repeats, each comprising five transmembrane helices. These repeats adopt asymmetric conformations, yielding an outward-facing occluded state. The crystal structure also revealed four putative ion-binding sites, but the occupancy and specificity thereof could not be conclusively established. Here, we use molecular-dynamics simulations and free-energy calculations to identify the ion configuration that best corresponds to the crystallographic data and that is also thermodynamically optimal. In this most probable configuration, three Na + ions occupy the so-called S ext , S Ca , and S int sites, whereas the S mid site is occupied by one water molecule and one H + , which protonates an adjacent aspartate side chain (D240). Experimental measurements of Na + /Ca 2+ and Ca 2+ /Ca 2+ exchange by wild-type and mutagenized NCX_Mj confirm that transport of both Na + and Ca 2+ requires protonation of D240, and that this side chain does not coordinate either ion at S mid . These results imply that the ion exchange stoichiometry of NCX_Mj is 3:1 and that translocation of Na + across the membrane is electrogenic, whereas transport of Ca 2+ is not. Altogether, these findings provide the basis for further experimental and computational studies of the conformational mechanism of this exchanger.secondary transporters | membrane antiporters | ion specificity | CaCA superfamily | molecular-dynamics simulations C a 2+ signals control a variety of cellular processes essential for the basic function of multiple organs. In cardiac cells, for example, Ca 2+ release from the sarcoplasmic reticulum is a necessary step for heart contraction, whereas Ca 2+ extrusion from the cell is required for cardiac relaxation. These fluctuations in the cytosolic Ca 2+ concentration underlie the initiation of the heartbeat (1, 2). Na + /Ca 2+ exchangers (NCXs) play a central role in the homeostasis of cellular Ca 2+ (3-5). These integral membrane proteins are ubiquitous in many types of tissues including the heart, brain, and kidney (4), and consequently their dysfunction is associated with numerous human pathologies such as cardiac arrhythmia, hypertension, skeletal muscle dystrophy, and postischemic brain damage (5). NCXs facilitate the translocation of either Ca 2+ or Na + across the membrane; thus, they can harness a transmembrane sodium motive force to energize Ca 2+ transport against a concentration gradient. For example, the cardiac exchanger NCX1 mediates the extrusion of intracellular Ca 2+ driven by a Na + transmembrane gradient maintained by the Na ...

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Section: Simulations Identify Three Ion Configurations Similarly Consmentioning

confidence: 77%

Sodium recognition by the Na ⁺ /Ca ²⁺ exchanger in the outward-facing conformation

Marinelli

Almagor

Hiller

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

124

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“…In LeuT, both substrates and noncompetitive inhibitors bind at the same site, which is located more extracellularly compared with the primary substrate binding site (reviewed in 123). Crystal structures of Na + (H + )-Ca 2+ exchangers in outward-and inward-facing conformations have shown that they, like LeuT fold transporters, are also made up of two structurally inverted repeats of five TM segments that come together to form two distinct bundles, referred to as a gate and core domain (124,125). It has been proposed that the gate domain, which is made of TM1 and TM6, rocks around the eight-TM-segment ion-binding core domain.…”

Section: Diversity In Local Gating and Global Rocking-bundle Movementsmentioning

confidence: 99%

Shared Molecular Mechanisms of Membrane Transporters

Drew

Boudker

2016

Annu. Rev. Biochem.

440

482

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The determination of the crystal structures of small-molecule transporters has shed light on the conformational changes that take place during structural isomerization from outward- to inward-facing states. Rather than using a simple rocking movement of two bundles around a central substrate-binding site, it has become clear that even the most simplistic transporters utilize rearrangements of nonrigid bodies. In the most dramatic cases, one bundle is fixed while the other, structurally divergent, bundle carries the substrate some 18 Å across the membrane, which in this review is termed an elevator alternating-access mechanism. Here, we compare and contrast rocker-switch, rocking-bundle, and elevator alternating-access mechanisms to highlight shared features and novel refinements to the basic alternating-access model.

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“…A substantial recent advance within CAX research has been the determination of detailed high‐resolution crystal structures of CAX proteins from yeast ( Saccharomyces cerevisiae ; Waight et al . 2013), as well as from bacterial CAXs (Nishizawa et al . 2013; Wu et al .…”

Section: Phylogenetic Diversity But Structural Conservation Of Cax Prmentioning

confidence: 99%

CAX‐ing a wide net: Cation/H⁺ transporters in metal remediation and abiotic stress signalling

2016

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Cation/proton exchangers (CAXs) are a class of secondary energised ion transporter that are being implicated in an increasing range of cellular and physiological functions. CAXs are primarily Ca2+ efflux transporters that mediate the sequestration of Ca2+ from the cytosol, usually into the vacuole. Some CAX isoforms have broad substrate specificity, providing the ability to transport trace metal ions such as Mn2+ and Cd2+, as well as Ca2+. In recent years, genomic analyses have begun to uncover the expansion of CAXs within the green lineage and their presence within non‐plant species. Although there appears to be significant conservation in tertiary structure of CAX proteins, there is diversity in function of CAXs between species and individual isoforms. For example, in halophytic plants, CAXs have been recruited to play a role in salt tolerance, while in metal hyperaccumulator plants CAXs are implicated in cadmium transport and tolerance. CAX proteins are involved in various abiotic stress response pathways, in some cases as a modulator of cytosolic Ca2+ signalling, but in some situations there is evidence of CAXs acting as a pH regulator. The metal transport and abiotic stress tolerance functions of CAXs make them attractive targets for biotechnology, whether to provide mineral nutrient biofortification or toxic metal bioremediation. The study of non‐plant CAXs may also provide insight into both conserved and novel transport mechanisms and functions.

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Structural basis for alternating access of a eukaryotic calcium/proton exchanger

Cited by 94 publications

References 54 publications

Sodium recognition by the Na ⁺ /Ca ²⁺ exchanger in the outward-facing conformation

Sodium recognition by the Na ⁺ /Ca ²⁺ exchanger in the outward-facing conformation

Shared Molecular Mechanisms of Membrane Transporters

CAX‐ing a wide net: Cation/H⁺ transporters in metal remediation and abiotic stress signalling

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Structural basis for alternating access of a eukaryotic calcium/proton exchanger

Cited by 94 publications

References 54 publications

Sodium recognition by the Na + /Ca 2+ exchanger in the outward-facing conformation

Sodium recognition by the Na + /Ca 2+ exchanger in the outward-facing conformation

Shared Molecular Mechanisms of Membrane Transporters

CAX‐ing a wide net: Cation/H+ transporters in metal remediation and abiotic stress signalling

Contact Info

Product

Resources

About

Sodium recognition by the Na ⁺ /Ca ²⁺ exchanger in the outward-facing conformation

Sodium recognition by the Na ⁺ /Ca ²⁺ exchanger in the outward-facing conformation

CAX‐ing a wide net: Cation/H⁺ transporters in metal remediation and abiotic stress signalling