%0 Journal Article
%T Structural and energetic analysis of metastable intermediate states in the E1P-E2P transition of Ca2+-ATPase.
%A Kobayashi C
%A Matsunaga Y
%A Jung J
%A Sugita Y
%J Proc Natl Acad Sci U S A
%V 118
%N 40
%D 10 2021 5
%M 34593638
%F 12.779
%R 10.1073/pnas.2105507118
%X Sarcoplasmic reticulum (SR) Ca2+-ATPase transports two Ca2+ ions from the cytoplasm to the SR lumen against a large concentration gradient. X-ray crystallography has revealed the atomic structures of the protein before and after the dissociation of Ca2+, while biochemical studies have suggested the existence of intermediate states in the transition between E1P⋅ADP⋅2Ca2+ and E2P. Here, we explore the pathway and free energy profile of the transition using atomistic molecular dynamics simulations with the mean-force string method and umbrella sampling. The simulations suggest that a series of structural changes accompany the ordered dissociation of ADP, the A-domain rotation, and the rearrangement of the transmembrane (TM) helices. The luminal gate then opens to release Ca2+ ions toward the SR lumen. Intermediate structures on the pathway are stabilized by transient sidechain interactions between the A- and P-domains. Lipid molecules between TM helices play a key role in the stabilization. Free energy profiles of the transition assuming different protonation states suggest rapid exchanges between Ca2+ ions and protons when the Ca2+ ions are released toward the SR lumen.