Structural dynamics of calmodulin-ryanodine receptor interactions: electron paramagnetic resonance using stereospecific spin labels

Abstract We have used electron paramagnetic resonance, with rigid and stereospecific spin labels, to resolve structural states in calmodulin (CaM), as affected by binding of Ca and a CaM-binding peptide (RyRp) derived from the ryanodine receptor (RyR), the Ca channel that triggers muscle contraction...

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Autores principales:	Cheng Her, Andrew R. Thompson, Christine B. Karim, David D. Thomas
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2018
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/e18d0469e22a4eb1a83261faac5a55bf
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spelling	oai:doaj.org-article:e18d0469e22a4eb1a83261faac5a55bf2021-12-02T15:08:18ZStructural dynamics of calmodulin-ryanodine receptor interactions: electron paramagnetic resonance using stereospecific spin labels10.1038/s41598-018-29064-82045-2322https://doaj.org/article/e18d0469e22a4eb1a83261faac5a55bf2018-07-01T00:00:00Zhttps://doi.org/10.1038/s41598-018-29064-8https://doaj.org/toc/2045-2322Abstract We have used electron paramagnetic resonance, with rigid and stereospecific spin labels, to resolve structural states in calmodulin (CaM), as affected by binding of Ca and a CaM-binding peptide (RyRp) derived from the ryanodine receptor (RyR), the Ca channel that triggers muscle contraction. CaM mutants containing a pair of cysteines in the N-lobe and/or C-lobe were engineered and labeled with a stereospecifically bound bifunctional spin label (BSL). RyRp was synthesized with and without TOAC (a stereospecifically attached spin-labeled amino acid) substituted for a single amino acid near the N-terminus. Intramolecular DEER distance measurements of doubly-labeled BSL-CaM revealed that CaM exists in dynamic equilibrium among multiple states, consistent with open, closed, and compact structural models. Addition of RyRp shifted the equilibrium partially toward the compact state in the absence of Ca, and completely toward the compact state in the presence of Ca, supporting a conformational selection model. Inter-protein distance measurements show that Ca stabilizes the compact state primarily by inducing ordered binding of the CaM N-lobe to RyRp, while only slightly affecting the C-lobe. The results provide insight into the structural mechanism of CaM-mediated RyR regulation, while demonstrating the power of using two types of rigidly and stereospecifically bound spin labels.Cheng HerAndrew R. ThompsonChristine B. KarimDavid D. ThomasNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 8, Iss 1, Pp 1-11 (2018)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Cheng Her Andrew R. Thompson Christine B. Karim David D. Thomas Structural dynamics of calmodulin-ryanodine receptor interactions: electron paramagnetic resonance using stereospecific spin labels
description	Abstract We have used electron paramagnetic resonance, with rigid and stereospecific spin labels, to resolve structural states in calmodulin (CaM), as affected by binding of Ca and a CaM-binding peptide (RyRp) derived from the ryanodine receptor (RyR), the Ca channel that triggers muscle contraction. CaM mutants containing a pair of cysteines in the N-lobe and/or C-lobe were engineered and labeled with a stereospecifically bound bifunctional spin label (BSL). RyRp was synthesized with and without TOAC (a stereospecifically attached spin-labeled amino acid) substituted for a single amino acid near the N-terminus. Intramolecular DEER distance measurements of doubly-labeled BSL-CaM revealed that CaM exists in dynamic equilibrium among multiple states, consistent with open, closed, and compact structural models. Addition of RyRp shifted the equilibrium partially toward the compact state in the absence of Ca, and completely toward the compact state in the presence of Ca, supporting a conformational selection model. Inter-protein distance measurements show that Ca stabilizes the compact state primarily by inducing ordered binding of the CaM N-lobe to RyRp, while only slightly affecting the C-lobe. The results provide insight into the structural mechanism of CaM-mediated RyR regulation, while demonstrating the power of using two types of rigidly and stereospecifically bound spin labels.
format	article
author	Cheng Her Andrew R. Thompson Christine B. Karim David D. Thomas
author_facet	Cheng Her Andrew R. Thompson Christine B. Karim David D. Thomas
author_sort	Cheng Her
title	Structural dynamics of calmodulin-ryanodine receptor interactions: electron paramagnetic resonance using stereospecific spin labels
title_short	Structural dynamics of calmodulin-ryanodine receptor interactions: electron paramagnetic resonance using stereospecific spin labels
title_full	Structural dynamics of calmodulin-ryanodine receptor interactions: electron paramagnetic resonance using stereospecific spin labels
title_fullStr	Structural dynamics of calmodulin-ryanodine receptor interactions: electron paramagnetic resonance using stereospecific spin labels
title_full_unstemmed	Structural dynamics of calmodulin-ryanodine receptor interactions: electron paramagnetic resonance using stereospecific spin labels
title_sort	structural dynamics of calmodulin-ryanodine receptor interactions: electron paramagnetic resonance using stereospecific spin labels
publisher	Nature Portfolio
publishDate	2018
url	https://doaj.org/article/e18d0469e22a4eb1a83261faac5a55bf
work_keys_str_mv	AT chengher structuraldynamicsofcalmodulinryanodinereceptorinteractionselectronparamagneticresonanceusingstereospecificspinlabels AT andrewrthompson structuraldynamicsofcalmodulinryanodinereceptorinteractionselectronparamagneticresonanceusingstereospecificspinlabels AT christinebkarim structuraldynamicsofcalmodulinryanodinereceptorinteractionselectronparamagneticresonanceusingstereospecificspinlabels AT daviddthomas structuraldynamicsofcalmodulinryanodinereceptorinteractionselectronparamagneticresonanceusingstereospecificspinlabels
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Structural dynamics of calmodulin-ryanodine receptor interactions: electron paramagnetic resonance using stereospecific spin labels

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