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Research Themes Membrane proteins

Membrane Proteome: Capturing Multiple Conformations

SBKB [doi:10.1038/sbkb.2012.111]
Featured Article - December 2012
Short description: Four novel crystallized conformers of the betaine/Na+ symporter BetP suggest a structural mechanism for molecular transport.

Alternating-access gating mechanism in BetP. 1

Among ubiquitous membrane transporters, which mediate the transport of a variety of solutes (including ions, neurotransmitters, nutrients and drugs) across lipid membranes, the secondary bacterial betaine transporter BetP is unique in functioning both as an osmosensor and osmoregulator. BetP senses hyperosmotic stress, specifically cytoplasmic K+ concentrations, and imports one molecule of choline/glycine betaine while taking up two Na+ ions. BetP forms a trimer, and each monomer adopts a LeuT-like fold of two inverted structural repeats with auxiliary terminal domains that mediate osmosensing and transport activity regulation.

Decades ago, transport was hypothesized to occur via an alternating access mechanism with isomerization between an outward-facing state with a substrate-binding vestibule accessible from the extracytoplasmic side, an optional occluded state in which the substrate is blocked from exiting on either side, and an inward-facing state with the binding site accessible from the cytoplasm. To a large extent, this model was confirmed by disparate structural “snapshots,” but no unified mechanism could be advanced without the capture of a single transporter in all isomerization states. Ziegler and colleagues now report the determination of two new BetP asymmetric trimeric crystal structures in the presence of betaine (PDB 4AIN) and choline (PDB 4DOJ), in which no less than five different conformers can be observed at once.

The study demonstrates not only how BetP switches from the substrate-free outward open (Ce) to inward-open (Ci) state, but also to the previously unobserved and transient closed substrate-free (Cc), closed-substrate-bound, inward-open betaine-bound, and inward-open choline-bound conformations. The authors attribute Ce/Cc/Ci transitions to a combination of rigid body rocking within a central bundle domain coupled to flexing of symmetry-related helices, and betaine binding to a tryptophan prism. Periplasmic residues have the greatest effect on betaine affinity, explaining its vectorial transport. The structures also illuminate a second aspect of molecular transport — the local opening and closing of the periplasmic and cytoplasmic gates — as well as the plasticity of the elements that coordinate sodium ions. Although the limited resolution (below 3Å) of the structures did not allow the unequivocal identification of the Na+ ions, recent biochemical and computational studies by Zhafizov and colleagues have given experimental support to the two proposed Na+-binding sites in BetP, thus paving the way to more targeted studies of this widespread family of proteins.

Alexandra Deaconescu

References

  1. C. Perez et al. Alternating-access mechanism in conformationally asymmetric trimers of the betaine transporter BetP.
    Nature 490, 126-130 (2 September 2012). doi:10.1038/nature11403

  2. K Khafizov et al. Investigation of the sodium-binding sites in the sodium-coupled betaine transporter BetP.
    Proc Natl Acad Sci USA 109, E3035-44 (30 October 2012). doi:10.1073/pnas.1209039109

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