Dynamics of P-type ATPase transport cycle revealed by single-molecule FRET

TY - JOUR

T1 - Dynamics of P-type ATPase transport cycle revealed by single-molecule FRET

AU - Dyla, Mateusz

AU - Terry, Daniel

AU - Kjærgaard, Magnus

AU - Sørensen, Thomas Lykke-Møller

AU - Andersen, Jacob Lauwring

AU - Andersen, Jens Peter

AU - Knudsen, Charlotte Rohde

AU - Altman, Roger

AU - Nissen, Poul

AU - Blanchard, Scott

PY - 2017/11/16

Y1 - 2017/11/16

N2 - Phosphorylation-type (P-type) ATPases are ubiquitous primary transporters that pump cations across cell membranes through the formation and breakdown of a phosphoenzyme intermediate. Structural investigations suggest that the transport mechanism is defined by conformational changes in the cytoplasmic domains of the protein that are allosterically coupled to transmembrane helices so as to expose ion binding sites to alternate sides of the membrane. Here, we have used single-molecule fluorescence resonance energy transfer to directly observe conformational changes associated with the functional transitions in the Listeria monocytogenes Ca 2+ -ATPase (LMCA1), an orthologue of eukaryotic Ca 2+ -ATPases. We identify key intermediates with no known crystal structures and show that Ca 2+ efflux by LMCA1 is rate-limited by phosphoenzyme formation. The transport process involves reversible steps and an irreversible step that follows release of ADP and extracellular release of Ca 2+.

AB - Phosphorylation-type (P-type) ATPases are ubiquitous primary transporters that pump cations across cell membranes through the formation and breakdown of a phosphoenzyme intermediate. Structural investigations suggest that the transport mechanism is defined by conformational changes in the cytoplasmic domains of the protein that are allosterically coupled to transmembrane helices so as to expose ion binding sites to alternate sides of the membrane. Here, we have used single-molecule fluorescence resonance energy transfer to directly observe conformational changes associated with the functional transitions in the Listeria monocytogenes Ca 2+ -ATPase (LMCA1), an orthologue of eukaryotic Ca 2+ -ATPases. We identify key intermediates with no known crystal structures and show that Ca 2+ efflux by LMCA1 is rate-limited by phosphoenzyme formation. The transport process involves reversible steps and an irreversible step that follows release of ADP and extracellular release of Ca 2+.

UR - http://www.scopus.com/inward/record.url?scp=85034453753&partnerID=8YFLogxK

U2 - 10.1038/nature24296

DO - 10.1038/nature24296

M3 - Journal article

C2 - 29144454

SN - 0028-0836

VL - 551

SP - 346

EP - 351

JO - Nature

JF - Nature

IS - 7680

ER -