Format

Send to

Choose Destination
Biophys J. 2016 Sep 20;111(6):1192-1202. doi: 10.1016/j.bpj.2016.08.005.

Cardiac Calcium ATPase Dimerization Measured by Cross-Linking and Fluorescence Energy Transfer.

Author information

1
Cell and Molecular Physiology, Loyola University Chicago, Chicago, Illinois.
2
Cell and Molecular Physiology, Loyola University Chicago, Chicago, Illinois. Electronic address: srobia@luc.edu.

Abstract

The cardiac sarco/endoplasmic reticulum calcium ATPase (SERCA) establishes the intracellular calcium gradient across the sarcoplasmic reticulum membrane. It has been proposed that SERCA forms homooligomers that increase the catalytic rate of calcium transport. We investigated SERCA dimerization in rabbit left ventricular myocytes using a photoactivatable cross-linker. Western blotting of cross-linked SERCA revealed higher-molecular-weight species consistent with SERCA oligomerization. Fluorescence resonance energy transfer measurements in cells transiently transfected with fluorescently labeled SERCA2a revealed that SERCA readily forms homodimers. These dimers formed in the absence or presence of the SERCA regulatory partner, phospholamban (PLB) and were unaltered by PLB phosphorylation or changes in calcium or ATP. Fluorescence lifetime data are compatible with a model in which PLB interacts with a SERCA homodimer in a stoichiometry of 1:2. Together, these results suggest that SERCA forms constitutive homodimers in live cells and that dimer formation is not modulated by SERCA conformational poise, PLB binding, or PLB phosphorylation.

PMID:
27653478
PMCID:
PMC5034344
DOI:
10.1016/j.bpj.2016.08.005
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Elsevier Science Icon for PubMed Central
Loading ...
Support Center