S100A1 protein does not compete with calmodulin for ryanodine receptor binding but structurally alters the ryanodine receptoro·calmodulin complex

Robyn T. Rebbeck, Florentin R. Nitu, David Rohde, Patrick Most, Donald M Bers, David D. Thomas, Razvan L. Cornea

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

S100A1 has been suggested as a therapeutic agent to enhance myocyte Ca2+ cycling in heart failure, but its molecular mode of action is poorly understood. Using FRET, we tested the hypothesis that S100A1 directly competes with calmodulin (CaM) for binding to intact, functional ryanodine receptors type I (RyR1) and II (RyR2) from skeletal and cardiac muscle, respectively. Our FRET readout provides an index of acceptor-labeled CaM binding near donor-labeled FKBP (FK506-binding protein 12.6) on the cytoplasmic domain of RyR in isolated sarcoplasmic reticulum vesicles. S100A1 (0.01-400 μM) partially inhibited FRET (i.e. CaM binding), with Ki > 10 μM, for both RyR1 and RyR2. The high [S100A1] required for partial effects on FRET indicates a lack of competition by S100A1 on CaM/RyR binding under normal physiological conditions. High-resolution analysis of time-resolved FRET detects two structural states of RyRbound CaM, which respond to [Ca2+] and are isoform-specific. The distribution of these structural states was perturbed only by high micromolar [S100A1], which promoted a shift of bound CaM to a lower FRET orientation (without altering the amount of CaM bound to RyR). Thus, high micromolar S100A1 does alter the CaM/RyR interaction, without involving competition. Nevertheless, submicromolar S100A1 can alter RyR function, an effect that is influenced by both [Ca2+] and [CaM]. We conclude that CaM and S100A1 can concurrently bind to and functionally modulate RyR1 and RyR2, but this does not involve direct competition at the RyR CaM binding site.

Original languageEnglish (US)
Pages (from-to)15896-15907
Number of pages12
JournalJournal of Biological Chemistry
Volume291
Issue number30
DOIs
StatePublished - Jul 22 2016

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Ryanodine
Ryanodine Receptor Calcium Release Channel
Calmodulin
S100A1 protein
Tacrolimus Binding Proteins
Sarcoplasmic Reticulum
Muscle Cells
Muscle
Myocardium
Protein Isoforms
Skeletal Muscle
Heart Failure
Binding Sites

ASJC Scopus subject areas

  • Biochemistry
  • Medicine(all)
  • Molecular Biology
  • Cell Biology

Cite this

S100A1 protein does not compete with calmodulin for ryanodine receptor binding but structurally alters the ryanodine receptoro·calmodulin complex. / Rebbeck, Robyn T.; Nitu, Florentin R.; Rohde, David; Most, Patrick; Bers, Donald M; Thomas, David D.; Cornea, Razvan L.

In: Journal of Biological Chemistry, Vol. 291, No. 30, 22.07.2016, p. 15896-15907.

Research output: Contribution to journalArticle

Rebbeck, Robyn T. ; Nitu, Florentin R. ; Rohde, David ; Most, Patrick ; Bers, Donald M ; Thomas, David D. ; Cornea, Razvan L. / S100A1 protein does not compete with calmodulin for ryanodine receptor binding but structurally alters the ryanodine receptoro·calmodulin complex. In: Journal of Biological Chemistry. 2016 ; Vol. 291, No. 30. pp. 15896-15907.
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abstract = "S100A1 has been suggested as a therapeutic agent to enhance myocyte Ca2+ cycling in heart failure, but its molecular mode of action is poorly understood. Using FRET, we tested the hypothesis that S100A1 directly competes with calmodulin (CaM) for binding to intact, functional ryanodine receptors type I (RyR1) and II (RyR2) from skeletal and cardiac muscle, respectively. Our FRET readout provides an index of acceptor-labeled CaM binding near donor-labeled FKBP (FK506-binding protein 12.6) on the cytoplasmic domain of RyR in isolated sarcoplasmic reticulum vesicles. S100A1 (0.01-400 μM) partially inhibited FRET (i.e. CaM binding), with Ki > 10 μM, for both RyR1 and RyR2. The high [S100A1] required for partial effects on FRET indicates a lack of competition by S100A1 on CaM/RyR binding under normal physiological conditions. High-resolution analysis of time-resolved FRET detects two structural states of RyRbound CaM, which respond to [Ca2+] and are isoform-specific. The distribution of these structural states was perturbed only by high micromolar [S100A1], which promoted a shift of bound CaM to a lower FRET orientation (without altering the amount of CaM bound to RyR). Thus, high micromolar S100A1 does alter the CaM/RyR interaction, without involving competition. Nevertheless, submicromolar S100A1 can alter RyR function, an effect that is influenced by both [Ca2+] and [CaM]. We conclude that CaM and S100A1 can concurrently bind to and functionally modulate RyR1 and RyR2, but this does not involve direct competition at the RyR CaM binding site.",
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AU - Thomas, David D.

AU - Cornea, Razvan L.

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AB - S100A1 has been suggested as a therapeutic agent to enhance myocyte Ca2+ cycling in heart failure, but its molecular mode of action is poorly understood. Using FRET, we tested the hypothesis that S100A1 directly competes with calmodulin (CaM) for binding to intact, functional ryanodine receptors type I (RyR1) and II (RyR2) from skeletal and cardiac muscle, respectively. Our FRET readout provides an index of acceptor-labeled CaM binding near donor-labeled FKBP (FK506-binding protein 12.6) on the cytoplasmic domain of RyR in isolated sarcoplasmic reticulum vesicles. S100A1 (0.01-400 μM) partially inhibited FRET (i.e. CaM binding), with Ki > 10 μM, for both RyR1 and RyR2. The high [S100A1] required for partial effects on FRET indicates a lack of competition by S100A1 on CaM/RyR binding under normal physiological conditions. High-resolution analysis of time-resolved FRET detects two structural states of RyRbound CaM, which respond to [Ca2+] and are isoform-specific. The distribution of these structural states was perturbed only by high micromolar [S100A1], which promoted a shift of bound CaM to a lower FRET orientation (without altering the amount of CaM bound to RyR). Thus, high micromolar S100A1 does alter the CaM/RyR interaction, without involving competition. Nevertheless, submicromolar S100A1 can alter RyR function, an effect that is influenced by both [Ca2+] and [CaM]. We conclude that CaM and S100A1 can concurrently bind to and functionally modulate RyR1 and RyR2, but this does not involve direct competition at the RyR CaM binding site.

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