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Alternating-access mechanism in conformationally asymmetric trimers of the betaine transporter BetP

Nature volume 490, pages 126–130 (2012)Cite this article

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Abstract

Betaine and Na+ symport has been extensively studied in the osmotically regulated transporter BetP from Corynebacterium glutamicum, a member of the betaine/choline/carnitine transporter family, which shares the conserved LeuT-like fold of two inverted structural repeats1. BetP adjusts its transport activity by sensing the cytoplasmic K+ concentration as a measure for hyperosmotic stress via the osmosensing carboxy-terminal domain2,3. BetP needs to be in a trimeric state for communication between individual protomers through several intratrimeric interaction sites4. Recently, crystal structures of inward-facing BetP trimers have contributed to our understanding of activity regulation on a molecular level5,6. Here we report new crystal structures, which reveal two conformationally asymmetric BetP trimers7, capturing among them three distinct transport states. We observe a total of four new conformations at once: an outward-open apo and an outward-occluded apo state, and two closed transition states—one in complex with betaine and one substrate-free. On the basis of these new structures, we identified local and global conformational changes in BetP that underlie the molecular transport mechanism, which partially resemble structural changes observed in other sodium-coupled LeuT-like fold transporters, but show differences we attribute to the osmolytic nature of betaine, the exclusive substrate specificity and the regulatory properties of BetP.

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Figure 1: Conformational states and substrate-binding sites observed in BetP and BetP(G153D) asymmetric trimers.
Figure 2: Conformational changes during the alternating-access cycle.
Figure 3: Opening and closing of the periplasmic and cytoplasmic gates in BetP.
Figure 4: Conformational changes of the sodium-binding sites.

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Primary accessions

Protein Data Bank

Data deposits

Coordinates and structure factors for the structures presented here have been deposited in the Protein Data Bank under accession 4DOJ and 4AIN.

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Acknowledgements

We thank L. Forrest and R. Krämer for careful reading of the manuscript and suggestions and discussions. This work was supported by the International Max-Planck Research School (C.K. and C.P.), and by the DFG (German Research Foundation), Collaborative Research Center 807 “Transport and Communication across Biological Membranes” (C.Z.).

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Author notes

  1. Camilo Perez and Caroline Koshy: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Structural Biology, Max-Planck Institute of Biophysics, D-60438 Frankfurt am Main, Germany

    Camilo Perez, Caroline Koshy, Özkan Yildiz & Christine Ziegler

  2. Institute of Biophysics and Biophysical Chemistry, University of Regensburg, D-93053 Regensburg, Germany,

    Christine Ziegler

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  1. Camilo Perez
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  2. Caroline Koshy
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Contributions

C.P. performed mutations, activity and binding measurements in cells and proteoliposomes, crystallization, collection and processing of data for PDB accession 4DOJ; C.K. performed crystallization for PDB accession 4AIN; C.K. and Ö.Y. performed collection and processing of data for PDB accession 4AIN; C.P., C.K. and C.Z. analysed the data; C.Z. directed the research; and C.P. and C.Z. wrote the manuscript.

Corresponding author

Correspondence to Christine Ziegler.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains Supplementary Methods, a Supplementary Discussion, Supplementary Tables 1-3, Supplementary Figures 1-10, a legend for Supplementary Movie 1 and Supplementary References. (PDF 6018 kb)

Supplementary Movie 1

The movie represents structural morphs of the conformational changes in BetP during transition from the outward-open to closed and inward-open states – see Supplementary Information file for full legend. (MP4 30132 kb)

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Perez, C., Koshy, C., Yildiz, Ö. et al. Alternating-access mechanism in conformationally asymmetric trimers of the betaine transporter BetP. Nature 490, 126–130 (2012). https://doi.org/10.1038/nature11403

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