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Unsynchronised subunit motion in single trimeric sodium-coupled aspartate transporters

Journal Article


Abstract


  • Excitatory amino acid transporters (EAATs) are secondary transport proteins that mediate the uptake of glutamate and other amino acids1. EAATs fulfil an important role in neuronal signal transmission by clearing the excitatory neurotransmitters from the synaptic cleft after depolarization of the postsynaptic neuron. An intensively studied model system for understanding the transport mechanism of EAATs is the archaeal aspartate transporter GltPh2, 3, 4, 5, 6. Each subunit in the homotrimeric GltPh supports the coupled translocation of one aspartate molecule and three Na+ ions2 as well as an uncoupled flux of Cl− ions7. Recent crystal structures of GltPh3, 5, 6, 8 revealed three possible conformations for the subunits, but it is unclear whether the motions of individual subunits are coordinated to support transport. Here, we report the direct observation of conformational dynamics in individual GltPh trimers embedded in the membrane by applying single-molecule fluorescence resonance energy transfer (FRET). By analysing the transporters in a lipid bilayer instead of commonly used detergent micelles, we achieve conditions that approximate the physiologically relevant ones. From the kinetics of FRET level transitions we conclude that the three GltPh subunits undergo conformational changes stochastically and independently of each other.

Authors


  •   Erkens, Guus B. (external author)
  •   Hanelt, Inga (external author)
  •   Goudsmits, Joris MH. (external author)
  •   Slotboom, Dirk J. (external author)
  •   van Oijen, Antoine M.

Publication Date


  • 2013

Published In


Citation


  • Erkens, G. B., Hanelt, I., Goudsmits, J. M. H., Slotboom, D. J. & van Oijen, A. M. (2013). Unsynchronised subunit motion in single trimeric sodium-coupled aspartate transporters. Nature, 502 (7469), 119-123.

Scopus Eid


  • 2-s2.0-84885606528

Ro Metadata Url


  • http://ro.uow.edu.au/smhpapers/2157

Has Global Citation Frequency


Number Of Pages


  • 4

Start Page


  • 119

End Page


  • 123

Volume


  • 502

Issue


  • 7469

Place Of Publication


  • United Kingdom

Abstract


  • Excitatory amino acid transporters (EAATs) are secondary transport proteins that mediate the uptake of glutamate and other amino acids1. EAATs fulfil an important role in neuronal signal transmission by clearing the excitatory neurotransmitters from the synaptic cleft after depolarization of the postsynaptic neuron. An intensively studied model system for understanding the transport mechanism of EAATs is the archaeal aspartate transporter GltPh2, 3, 4, 5, 6. Each subunit in the homotrimeric GltPh supports the coupled translocation of one aspartate molecule and three Na+ ions2 as well as an uncoupled flux of Cl− ions7. Recent crystal structures of GltPh3, 5, 6, 8 revealed three possible conformations for the subunits, but it is unclear whether the motions of individual subunits are coordinated to support transport. Here, we report the direct observation of conformational dynamics in individual GltPh trimers embedded in the membrane by applying single-molecule fluorescence resonance energy transfer (FRET). By analysing the transporters in a lipid bilayer instead of commonly used detergent micelles, we achieve conditions that approximate the physiologically relevant ones. From the kinetics of FRET level transitions we conclude that the three GltPh subunits undergo conformational changes stochastically and independently of each other.

Authors


  •   Erkens, Guus B. (external author)
  •   Hanelt, Inga (external author)
  •   Goudsmits, Joris MH. (external author)
  •   Slotboom, Dirk J. (external author)
  •   van Oijen, Antoine M.

Publication Date


  • 2013

Published In


Citation


  • Erkens, G. B., Hanelt, I., Goudsmits, J. M. H., Slotboom, D. J. & van Oijen, A. M. (2013). Unsynchronised subunit motion in single trimeric sodium-coupled aspartate transporters. Nature, 502 (7469), 119-123.

Scopus Eid


  • 2-s2.0-84885606528

Ro Metadata Url


  • http://ro.uow.edu.au/smhpapers/2157

Has Global Citation Frequency


Number Of Pages


  • 4

Start Page


  • 119

End Page


  • 123

Volume


  • 502

Issue


  • 7469

Place Of Publication


  • United Kingdom