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Single-molecule visualization of conformational changes and substrate transport in the vitamin B12ABC importer BtuCD-F

Journal Article


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Abstract


  • ATP-binding cassette (ABC) transporters form the largest class of active membrane transport proteins. Binding and hydrolysis of ATP by their highly conserved nucleotide-binding domains drive conformational changes of the complex that mediate transport of substrate across the membrane. The vitamin B 12 importer BtuCD-F in Escherichia coli is an extensively studied model system. The periplasmic soluble binding protein BtuF binds the ligand; the transmembrane and ATPase domains BtuCD mediate translocation. Here we report the direct observation at the single-molecule level of ATP, vitamin B 12 and BtuF-induced events in the transporter complex embedded in liposomes. Single-molecule fluorescence imaging techniques reveal that membrane-embedded BtuCD forms a stable complex with BtuF, regardless of the presence of ATP and vitamin B 12 . We observe that a vitamin B 12 molecule remains bound to the complex for tens of seconds, during which several ATP hydrolysis cycles can take place, before it is being transported across the membrane.

Authors


  •   Goudsmits, Joris MH. (external author)
  •   Slotboom, Dirk J. (external author)
  •   van Oijen, Antoine M.

Publication Date


  • 2017

Citation


  • Goudsmits, J. M. H., Slotboom, D. Jan. & van Oijen, A. M. (2017). Single-molecule visualization of conformational changes and substrate transport in the vitamin B12ABC importer BtuCD-F. Nature Communications, 8 (1), 1652-1-1652-10.

Scopus Eid


  • 2-s2.0-85034759100

Ro Full-text Url


  • http://ro.uow.edu.au/cgi/viewcontent.cgi?article=6162&context=smhpapers

Ro Metadata Url


  • http://ro.uow.edu.au/smhpapers/5109

Start Page


  • 1652-1

End Page


  • 1652-10

Volume


  • 8

Issue


  • 1

Place Of Publication


  • United Kingdom

Abstract


  • ATP-binding cassette (ABC) transporters form the largest class of active membrane transport proteins. Binding and hydrolysis of ATP by their highly conserved nucleotide-binding domains drive conformational changes of the complex that mediate transport of substrate across the membrane. The vitamin B 12 importer BtuCD-F in Escherichia coli is an extensively studied model system. The periplasmic soluble binding protein BtuF binds the ligand; the transmembrane and ATPase domains BtuCD mediate translocation. Here we report the direct observation at the single-molecule level of ATP, vitamin B 12 and BtuF-induced events in the transporter complex embedded in liposomes. Single-molecule fluorescence imaging techniques reveal that membrane-embedded BtuCD forms a stable complex with BtuF, regardless of the presence of ATP and vitamin B 12 . We observe that a vitamin B 12 molecule remains bound to the complex for tens of seconds, during which several ATP hydrolysis cycles can take place, before it is being transported across the membrane.

Authors


  •   Goudsmits, Joris MH. (external author)
  •   Slotboom, Dirk J. (external author)
  •   van Oijen, Antoine M.

Publication Date


  • 2017

Citation


  • Goudsmits, J. M. H., Slotboom, D. Jan. & van Oijen, A. M. (2017). Single-molecule visualization of conformational changes and substrate transport in the vitamin B12ABC importer BtuCD-F. Nature Communications, 8 (1), 1652-1-1652-10.

Scopus Eid


  • 2-s2.0-85034759100

Ro Full-text Url


  • http://ro.uow.edu.au/cgi/viewcontent.cgi?article=6162&context=smhpapers

Ro Metadata Url


  • http://ro.uow.edu.au/smhpapers/5109

Start Page


  • 1652-1

End Page


  • 1652-10

Volume


  • 8

Issue


  • 1

Place Of Publication


  • United Kingdom