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CFTR channel opening by ATP-driven tight dimerization of its nucleotide-binding domains

Nature volume 433, pages 876–880 (2005)Cite this article

Abstract

ABC (ATP-binding cassette) proteins constitute a large family of membrane proteins that actively transport a broad range of substrates. Cystic fibrosis transmembrane conductance regulator (CFTR), the protein dysfunctional in cystic fibrosis, is unique among ABC proteins in that its transmembrane domains comprise an ion channel. Opening and closing of the pore have been linked to ATP binding and hydrolysis at CFTR's two nucleotide-binding domains, NBD1 and NBD2 (see, for example, refs 1, 2). Isolated NBDs of prokaryotic ABC proteins dimerize upon binding ATP, and hydrolysis of the ATP causes dimer dissociation3,4,5. Here, using single-channel recording methods on intact CFTR molecules, we directly follow opening and closing of the channel gates, and relate these occurrences to ATP-mediated events in the NBDs. We find that energetic coupling6 between two CFTR residues, expected to lie on opposite sides of its predicted NBD1–NBD2 dimer interface, changes in concert with channel gating status. The two monitored side chains are independent of each other in closed channels but become coupled as the channels open. The results directly link ATP-driven tight dimerization of CFTR's cytoplasmic nucleotide-binding domains to opening of the ion channel in the transmembrane domains. This establishes a molecular mechanism, involving dynamic restructuring of the NBD dimer interface, that is probably common to all members of the ABC protein superfamily.

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Figure 1: Open CFTR channels correspond to dimerized NBDs.
Figure 2: Statistical coupling analysis and electrophysiological recordings position CFTR's Arg 555 in the composite NBD2 catalytic site.
Figure 3: Arg 555 and Thr 1246 are not energetically coupled in channel closed states.
Figure 4: Energetic coupling between Arg 555 and Thr 1246 accompanies channel opening.

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Acknowledgements

We thank L. Csanády and G. Szakács for discussion. The work was supported by an NIH grant to D.C.G.

Author information

Authors and Affiliations

  1. Laboratory of Cardiac/Membrane Physiology, The Rockefeller University, New York, New York, 10021, USA

    Paola Vergani & David C. Gadsby

  2. Laboratory of Molecular Neurobiology and Biophysics, The Rockefeller University, New York, New York, 10021, USA

    Steve W. Lockless

  3. Laboratory of Molecular and Cellular Neuroscience, The Rockefeller University, New York, New York, 10021, USA

    Angus C. Nairn

  4. Department of Psychiatry, Yale University, New Haven, Connecticut, 06519, USA

    Angus C. Nairn

Authors
  1. Paola Vergani
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  2. Steve W. Lockless
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  4. David C. Gadsby
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Corresponding authors

Correspondence to Paola Vergani or David C. Gadsby.

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Supplementary information

Supplementary Notes

Thermodynamic double mutant cycles. Derivation of equations used to quantify energetic coupling. (DOC 72 kb)

Supplementary Table 1

Kinetic Parameters of WT and mutant CFTR channels. Part A: kinetic parameters describing single-channel gating, obtained from patches containing a small number of channels (for example Figs 2c, 3b and 4b). Part B: Current decay time-constants, obtained from patches containing hundreds of channels (for example Fig. 1b). (DOC 23 kb)

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Vergani, P., Lockless, S., Nairn, A. et al. CFTR channel opening by ATP-driven tight dimerization of its nucleotide-binding domains. Nature 433, 876–880 (2005). https://doi.org/10.1038/nature03313

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