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Cardiac Calcium ATPase Dimerization Measured by Cross-Linking and Fluorescence Energy Transfer.
- Source :
-
Biophysical journal [Biophys J] 2016 Sep 20; Vol. 111 (6), pp. 1192-1202. - Publication Year :
- 2016
-
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.<br /> (Copyright © 2016 Biophysical Society. Published by Elsevier Inc. All rights reserved.)
- Subjects :
- Adenosine Triphosphate chemistry
Adenosine Triphosphate metabolism
Animals
Blotting, Western
Calcium chemistry
Calcium metabolism
Calcium-Binding Proteins chemistry
Calcium-Binding Proteins metabolism
Dogs
Fluorescence Resonance Energy Transfer
HEK293 Cells
Heart Ventricles chemistry
Heart Ventricles enzymology
Humans
Immunoprecipitation
Luminescent Proteins chemistry
Luminescent Proteins genetics
Luminescent Proteins metabolism
Models, Molecular
Muscle Cells chemistry
Muscle Cells enzymology
Mutation
Phosphorylation
Photobleaching
Protein Multimerization
Rabbits
Sarcoplasmic Reticulum Calcium-Transporting ATPases genetics
Sarcoplasmic Reticulum Calcium-Transporting ATPases metabolism
Sarcoplasmic Reticulum Calcium-Transporting ATPases chemistry
Subjects
Details
- Language :
- English
- ISSN :
- 1542-0086
- Volume :
- 111
- Issue :
- 6
- Database :
- MEDLINE
- Journal :
- Biophysical journal
- Publication Type :
- Academic Journal
- Accession number :
- 27653478
- Full Text :
- https://doi.org/10.1016/j.bpj.2016.08.005