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Design, function and structure of a monomeric ClC transporter


Channels and transporters of the ClC family cause the transmembrane movement of inorganic anions in service of a variety of biological tasks, from the unusual—the generation of the kilowatt pulses with which electric fish stun their prey—to the quotidian—the acidification of endosomes, vacuoles and lysosomes1. The homodimeric architecture of ClC proteins, initially inferred from single-molecule studies of an elasmobranch Cl channel2 and later confirmed by crystal structures of bacterial Cl/H+ antiporters3,4, is apparently universal. Moreover, the basic machinery that enables ion movement through these proteins—the aqueous pores for anion diffusion in the channels and the ion-coupling chambers that coordinate Cl and H+ antiport in the transporters—are contained wholly within each subunit of the homodimer. The near-normal function of a bacterial ClC transporter straitjacketed by covalent crosslinks across the dimer interface and the behaviour of a concatemeric human homologue argue that the transport cycle resides within each subunit and does not require rigid-body rearrangements between subunits5,6. However, this evidence is only inferential, and because examples are known in which quaternary rearrangements of extramembrane ClC domains that contribute to dimerization modulate transport activity7, we cannot declare as definitive a ‘parallel-pathways’ picture in which the homodimer consists of two single-subunit transporters operating independently. A strong prediction of such a view is that it should in principle be possible to obtain a monomeric ClC. Here we exploit the known structure of a ClC Cl/H+ exchanger, ClC-ec1 from Escherichia coli, to design mutants that destabilize the dimer interface while preserving both the structure and the transport function of individual subunits. The results demonstrate that the ClC subunit alone is the basic functional unit for transport and that cross-subunit interaction is not required for Cl/H+ exchange in ClC transporters.

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Figure 1: Structure and dimeric interface of ClC-ec1.
Figure 2: Behaviour of tryptophan mutants in detergent.
Figure 3: Monomeric ClC mutant in phospholipid membranes.
Figure 4: Crystal structure of the WW monomer.

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Primary accessions

Protein Data Bank

Data deposits

The atomic coordinates and structure factors of the WW (Ile201Trp/Ile422Trp) monomer have been deposited in the Protein Data Bank under accession code 3NMO.


  1. Jentsch, T. J. et al. Physiological functions of CLC Cl channels gleaned from human genetic disease and mouse models. Annu. Rev. Physiol. 67, 779–807 (2005)

    Article  CAS  Google Scholar 

  2. Middleton, R. E., Pheasant, D. J. & Miller, C. Reconstitution of detergent-solubilized Cl channels and analysis by concentrative uptake of 36Cl and planar lipid bilayers. Methods 6, 28–36 (1994)

    Article  CAS  Google Scholar 

  3. Dutzler, R. et al. X-ray structure of a ClC chloride channel at 3.0 Å reveals the molecular basis of anion selectivity. Nature 415, 287–294 (2002)

    Article  ADS  CAS  Google Scholar 

  4. Dutzler, R., Campbell, E. B. & MacKinnon, R. Gating the selectivity filter in ClC chloride channels. Science 300, 108–112 (2003)

    Article  ADS  CAS  Google Scholar 

  5. Nguitragool, W. & Miller, C. CLC Cl/H+ transporters constrained by covalent cross-linking. Proc. Natl Acad. Sci. USA 104, 20659–20665 (2007)

    Article  ADS  CAS  Google Scholar 

  6. Zdebik, A. A. et al. Determinants of anion-proton coupling in mammalian endosomal CLC proteins. J. Biol. Chem. 283, 4219–4227 (2008)

    Article  CAS  Google Scholar 

  7. Bykova, E. A. et al. Large movement in the C terminus of CLC-0 chloride channel during slow gating. Nature Struct. Mol. Biol. 13, 1115–1119 (2006)

    Article  CAS  Google Scholar 

  8. Fleming, K. G., Ackerman, A. L. & Engelman, D. A. The effect of point mutations on the free energy of transmembrane α-helix dimerization. J. Mol. Biol. 272, 266–275 (1997)

    Article  CAS  Google Scholar 

  9. MacKenzie, K. R. & Fleming, K. G. Association energetics of membrane spanning α-helices. Curr. Opin. Struct. Biol. 18, 412–419 (2008)

    Article  CAS  Google Scholar 

  10. Chen, L. et al. Energetics of ErbB1 transmembrane domain dimerization in lipid bilayers. Biophys. J. 96, 4622–4630 (2009)

    Article  ADS  CAS  Google Scholar 

  11. Yau, W. M. et al. The preference of tryptophan for membrane interfaces. Biochemistry 37, 14713–14718 (1998)

    Article  CAS  Google Scholar 

  12. Fang, Y., Kolmakova-Partensky, L. & Miller, C. A bacterial arginine-agmatine exchange transporter involved in extreme acid resistance. J. Biol. Chem. 282, 176–182 (2007)

    Article  CAS  Google Scholar 

  13. Maduke, M., Pheasant, D. J. & Miller, C. High-level expression, functional reconstitution, and quaternary structure of a prokaryotic ClC-type chloride channel. J. Gen. Physiol. 114, 713–722 (1999)

    Article  CAS  Google Scholar 

  14. Walden, M. et al. Uncoupling and turnover in a Cl/H+ exchange transporter. J. Gen. Physiol. 129, 317–329 (2007)

    Article  CAS  Google Scholar 

  15. Accardi, A. & Miller, C. Secondary active transport mediated by a prokaryotic homologue of ClC Cl- channels. Nature 427, 803–807 (2004)

    Article  ADS  CAS  Google Scholar 

  16. Miller, C. & Nguitragool, W. A provisional transport mechanism for a chloride channel-type Cl/H+ exchanger. Phil. Trans. R. Soc. B 364, 175–180 (2008)

    Article  Google Scholar 

  17. Elvington, S. M., Liu, C. W. & Maduke, M. C. Substrate-driven conformational changes in ClC-ec1 observed by fluorine NMR. EMBO J. 28, 3090–3102 (2009)

    Article  CAS  Google Scholar 

  18. Accardi, A. et al. Synergism between halide binding and proton transport in a CLC-type exchanger. J. Mol. Biol. 362, 691–699 (2006)

    Article  CAS  Google Scholar 

  19. Nguitragool, W. & Miller, C. Uncoupling of a CLC Cl/H+ exchange transporter by polyatomic anions. J. Mol. Biol. 362, 682–690 (2006)

    Article  CAS  Google Scholar 

  20. Fu, D. et al. Structure of a glycerol-conducting channel and the basis for its selectivity. Science 290, 481–486 (2000)

    Article  ADS  CAS  Google Scholar 

  21. Wang, Y. et al. Structure of the formate transporter FocA reveals a pentameric aquaporin-like channel. Nature 462, 467–472 (2009)

    Article  ADS  CAS  Google Scholar 

  22. Waight, A. B., Love, J. & Wang, D. N. Structure and mechanism of a pentameric formate channel. Nature Struct. Mol. Biol. 17, 31–37 (2010)

    Article  CAS  Google Scholar 

  23. Levin, E. J., Quick, M. & Zhou, M. Crystal structure of a bacterial homologue of the kidney urea transporter. Nature 462, 757–761 (2009)

    Article  ADS  CAS  Google Scholar 

  24. Khademi, S. et al. Mechanism of ammonia transport by Amt/MEP/Rh: structure of AmtB at 1.35 Å. Science 305, 1587–1594 (2004)

    Article  ADS  CAS  Google Scholar 

  25. Zheng, L. et al. The mechanism of ammonia transport based on the crystal structure of AmtB of Escherichia coli . Proc. Natl Acad. Sci. USA 101, 17090–17095 (2004)

    Article  ADS  CAS  Google Scholar 

  26. Cowan, S. W. et al. Crystal structures explain functional properties of two E. coli porins. Nature 358, 727–733 (1992)

    Article  ADS  CAS  Google Scholar 

  27. Shaffer, P. L. et al. Structure and mechanism of a Na+-independent amino acid transporter. Science 325, 1010–1014 (2009)

    Article  ADS  CAS  Google Scholar 

  28. Theobald, D. L. & Miller, C. Membrane transport proteins: surprises in structural sameness. Nature Struct. Mol. Biol. 17, 2–3 (2010)

    Article  CAS  Google Scholar 

  29. Reyes, N., Ginter, C. & Boudker, O. Transport mechanism of a bacterial homologue of glutamate transporters. Nature 462, 880–885 (2009)

    Article  ADS  CAS  Google Scholar 

  30. Metcalf, D. G. et al. Multiple approaches converge on the structure of the integrin αIIb/β3 transmembrane heterodimer. J. Mol. Biol. 392, 1087–1101 (2009)

    Article  CAS  Google Scholar 

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We are grateful to the scientists at beamline 8.2.1 of the Advanced Light Source, Lawrence Berkeley Laboratory, for much help and advice, to H. Jayaram for help with refinement, and to R. Sah and D. Theobald for comments on the manuscript.

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Experiments were designed by J.L.R. and C.M. and carried out by J.L.R. and L.K.-P., and the manuscript was written by all authors.

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Correspondence to Christopher Miller.

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The authors declare no competing financial interests.

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Robertson, J., Kolmakova-Partensky, L. & Miller, C. Design, function and structure of a monomeric ClC transporter. Nature 468, 844–847 (2010).

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