Redistribution of SERCA calcium pump conformers during intracellular calcium signaling

Olga N. Raguimova, Nikolai Smolin, Elisa Bovo, Siddharth Bhayani, Joseph M. Autry, Aleksey V. Zima, Seth L. Robia

Research output: Contribution to journalArticlepeer-review

21 Scopus citations

Abstract

The conformational changes of a calcium transport ATPase were investigated with molecular dynamics (MD) simulations as well as fluorescence resonance energy transfer (FRET) measurements to determine the significance of a discrete structural element for regulation of the conformational dynamics of the transport cycle. Previous MD simulations indicated that a loop in the cytosolic domain of the SERCA calcium transporter facilitates an open-to-closed structural transition. To investigate the significance of this structural element, we performed additional MD simulations and new biophysical measurements of SERCA structure and function. Rationally designed in silico mutations of three acidic residues of the loop decreased SERCA domain–domain contacts and increased domain– domain separation distances. Principal component analysis of MD simulations suggested decreased sampling of compact conformations upon N-loop mutagenesis. Deficits in headpiece structural dynamics were also detected by measuring intramolecular FRET of a Cer–YFP–SERCA construct (2-color SERCA). Compared with WT, the mutated 2-color SERCA shows a partial FRET response to calcium, whereas retaining full responsiveness to the inhibitor thapsigargin. Functional measurements showed that the mutated transporter still hydrolyzes ATP and transports calcium, but that maximal enzyme activity is reduced while maintaining similar calcium affinity. In live cells, calcium elevations resulted in concomitant FRET changes as the population of WT 2-color SERCA molecules redistributed among intermediates of the transport cycle. Our results provide novel insights on how the population of SERCA pumps responds to dynamic changes in intracellular calcium.

Original languageEnglish (US)
Pages (from-to)10843-10856
Number of pages14
JournalJournal of Biological Chemistry
Volume293
Issue number28
DOIs
StatePublished - Jul 13 2018

Bibliographical note

Funding Information:
Acknowledgments—We are grateful for helpful suggestions from Howard S. Young. This work was also supported by equipment and facilities provided by National Institute of Health “Loyola Research Computing Core” Grant 1G20RR030939. This work used the Extreme Science and Engineering Discovery Environment (XSEDE), which is supported by National Science Foundation Grant ACI-1548562, as well as Stampede and Stampede2 at the Texas Advanced Computing Center (TACC) through XSEDE allocation TG-MCB130108.

Funding Information:
This work was supported by the National Institutes of Health Grants HL092321 (to S. L. R) and HL130231 (to A. V. Z.) and the Loyola Cardiovas-cular Research Institute. The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the respon-sibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

Funding Information:
This work was supported by the National Institutes of Health Grants HL092321 (to S. L. R) and HL130231 (to A. V. Z.) and the Loyola Cardiovascular Research Institute. The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. We are grateful for helpful suggestions from Howard S. Young. This work was also supported by equipment and facilities provided by National Institute of Health “Loyola Research Computing Core” Grant 1G20RR030939. This work used the Extreme Science and Engineering Discovery Environment (XSEDE), which is supported by National Science Foundation Grant ACI-1548562, as well as Stampede and Stampede2 at the Texas Advanced Computing Center (TACC) through XSEDE allocation TG-MCB130108.

Publisher Copyright:
© 2018 Raguimova et al.

Fingerprint

Dive into the research topics of 'Redistribution of SERCA calcium pump conformers during intracellular calcium signaling'. Together they form a unique fingerprint.

Cite this