Ion and lipid orchestration of secondary active transport

Drew, David; Boudker, Olga

doi:10.1038/s41586-024-07062-3

Cited by 7 publications

(3 citation statements)

References 189 publications

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“…Another way to improve the structure-dynamic resolution of 19 F-NMR data (obtained in the present study) is to adopt new experimental approaches of 19 F-NMR (recently developed for a glutamate transporter homolog) [21,22,47,48] for the future analysis of nanodisc-reconstituted NCX_Mj. This new approach would involve the site-directed cys- 19 F-probe introduction at specific locations of the cytosolic and extracellular vestibules (as carried out in the present work), where a two-his-tag will be attached to a neighboring transmembrane helix (TM3 or TM8) to chelate Ni 2+ as a paramagnetic probe.…”

Section: Discussionmentioning

confidence: 99%

“…In contrast with ion channels, the "secondary active transporters" (antiporters, symporters, and uniporters) comply with the alternative access paradigm according to which the ligand (ion)-binding domain undergoes an alternative exposure to opposite sides of the membrane-so, successive transitions between the outward-facing (OF) and inwardfacing (IF) states accomplish the ion transport cycle [19][20][21][22]. Although the paradigm of alternative access is generally accepted, the structure-dynamic transitions along the transport cycle might differ in distinct sub-types of secondary active transporters, since the OF/IF swapping may occur either in the absence and/or presence of the ligand [20,23].…”

Section: Introductionmentioning

confidence: 99%

“…For example, in antiporters (like NCXs), the ligand (ion) interactions with respective transport site(s) are obligatory to induce the OF/IF swapping [20,23]. In contrast with antiporters, symporters and uniporters can perform the OF/IF shuttling either in the absence or presence of a ligand/ion [20][21][22][23]. The underlying structure-dynamic mechanisms remain unresolved even for transporters with known structure since available structural snapshots cannot provide a full picture of functionally important intermediates contributing to the transport cycle.…”

Section: Introductionmentioning

confidence: 99%

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19F-NMR Probing of Ion-Induced Conformational Changes in Detergent-Solubilized and Nanodisc-Reconstituted NCX_Mj

Nguyen,

Strauss,

Refaeli

et al. 2024

IJMS

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Consecutive interactions of 3Na+ or 1Ca2+ with the Na+/Ca2+ exchanger (NCX) result in an alternative exposure (access) of the cytosolic and extracellular vestibules to opposite sides of the membrane, where ion-induced transitions between the outward-facing (OF) and inward-facing (IF) conformational states drive a transport cycle. Here, we investigate sub-state populations of apo and ion-bound species in the OF and IF states by analyzing detergent-solubilized and nanodisc-reconstituted preparations of NCX_Mj with 19F-NMR. The 19F probe was covalently attached to the cysteine residues at entry locations of the cytosolic and extracellular vestibules. Multiple sub-states of apo and ion-bound species were observed in nanodisc-reconstituted (but not in detergent-solubilized) NCX_Mj, meaning that the lipid-membrane environment preconditions multiple sub-state populations toward the OF/IF swapping. Most importantly, ion-induced sub-state redistributions occur within each major (OF or IF) state, where sub-state interconversions may precondition the OF/IF swapping. In contrast with large changes in population redistributions, the sum of sub-state populations within each inherent state (OF or IF) remains nearly unchanged upon ion addition. The present findings allow the further elucidation of structure–dynamic modules underlying ion-induced conformational changes that determine a functional asymmetry of ion access/translocation at opposite sides of the membrane and ion transport rates concurring physiological demands.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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19F-NMR Probing of Ion-Induced Conformational Changes in Detergent-Solubilized and Nanodisc-Reconstituted NCX_Mj

Nguyen,

Strauss,

Refaeli

et al. 2024

IJMS

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Future opportunities in solute carrier structural biology

Newstead

2024

Nat Struct Mol Biol

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Molecular principles of redox-coupled sodium pumping of the ancient Rnf machinery

Kumar,

Roth,

Kim

et al. 2024

Preprint

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The Rnf complex is the primary respiratory enzyme of several anaerobic prokaryotes that transfers electrons from ferredoxin to NAD+and pumps sodium ions (Na+) across a membrane, powering ATP synthesis. Rnf is widespread in primordial organisms and the evolutionary predecessor of the Na+-pumping NADH-quinone oxidoreductase (Nqr)1. By running in reverse, Rnf reduces ferredoxin with NADH as reductant at the expense of the transmembrane electrochemical ion gradient and provides low potential electrons for nitrogenases as well as CO2reductases. Yet, the molecular principles that couple the long-range electron transfer to the Na+translocation across the membrane remain elusive. Here we resolve key functional states along the electron transfer pathway using redox-controlled cryo-electron microscopy (cryo-EM) that, in combination with biochemical functional assays and atomistic molecular simulations, provide key insight into the redox-driven Na+pumping mechanism. We show that the reduction of the unique membrane-embedded [2Fe2S] cluster in the vestibule between the RnfA/E subunits electrostatically attracts the sodium ions, and in turn, triggers aninward/outwardtransition with alternating membrane access driving the Na+pump and the reduction of NAD+. Our study unveils an ancient mechanism for redox-driven ion pumping, and provides key understanding of the fundamental principles governing energy conversion in biological systems.

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Ion and lipid orchestration of secondary active transport

Cited by 7 publications

References 189 publications

19F-NMR Probing of Ion-Induced Conformational Changes in Detergent-Solubilized and Nanodisc-Reconstituted NCX_Mj

19F-NMR Probing of Ion-Induced Conformational Changes in Detergent-Solubilized and Nanodisc-Reconstituted NCX_Mj

Future opportunities in solute carrier structural biology

Molecular principles of redox-coupled sodium pumping of the ancient Rnf machinery

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