Tracking Ca2+ ATPase intermediates in real time by x-ray solution scattering

Harsha Ravishankar, Martin Nors Pedersen*, Mattias Eklund, Aljona Sitsel, Chenge Li, Annette Duelli, Matteo Levantino, Michael Wulff, Andreas Barth, Claus Olesen, Poul Nissen, Magnus Andersson

*Corresponding author for this work

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaperJournal articleResearchpeer-review

1 Citation (Scopus)

Abstract

Sarco/endoplasmic reticulum Ca2+ ATPase (SERCA) transporters regulate calcium signaling by active calcium ion reuptake to internal stores. Structural transitions associated with transport have been characterized by x-ray crystallography, but critical intermediates involved in the accessibility switch across the membrane are missing. We combined time-resolved x-ray solution scattering (TR-XSS) experiments and molecular dynamics (MD) simulations for real-time tracking of concerted SERCA reaction cycle dynamics in the native membrane. The equilibrium [Ca2] E1 state before laser activation differed in the domain arrangement compared with crystal structures, and following laser-induced release of caged ATP, a 1.5-ms intermediate was formed that showed closure of the cytoplasmic domains typical of E1 states with bound Ca2+ and ATP. A subsequent 13-ms transient state showed a previously unresolved actuator (A) domain arrangement that exposed the ADP-binding site after phosphorylation. Hence, the obtained TR-XSS models determine the relative timing of so-far elusive domain rearrangements in a native environment.

Original languageEnglish
Article numbereaaz0981
JournalScience Advances
Volume6
Issue12
Number of pages10
ISSN2375-2548
DOIs
Publication statusPublished - Mar 2020

Keywords

  • CA2+-ATPASE
  • CALCIUM-TRANSPORT
  • CONFORMATIONAL-CHANGES
  • MEMBRANE
  • MOLECULAR-DYNAMICS
  • PHOSPHOENZYME
  • PUMP
  • SARCOPLASMIC-RETICULUM
  • SKELETAL-MUSCLE
  • STRUCTURAL DYNAMICS

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