1H, 15N, and 13C chemical shift assignments of the regulatory domain of human calcineurin

Dinesh K. Yadav; Sri Ramya Tata; John Hunt; Erik C. Cook; Trevor P. Creamer; Nicholas C. Fitzkee

doi:10.1007/s12104-017-9751-x

¹H, ¹⁵N, and ¹³C chemical shift assignments of the regulatory domain of human calcineurin

Dinesh K. Yadav, Sri Ramya Tata, John Hunt, Erik C. Cook, Trevor P. Creamer, Nicholas C. Fitzkee

Research output: Contribution to journal › Article › peer-review

3 Scopus citations

Abstract

Calcineurin (CaN) plays an important role in T-cell activation, cardiac system development and nervous system function. Previous studies have demonstrated that the regulatory domain (RD) of CaN binds calmodulin (CaM) towards the N-terminal end. Calcium-loaded CaM activates the serine/threonine phosphatase activity of CaN by binding to the RD, although the mechanistic details of this interaction remain unclear. It is thought that CaM binding at the RD displaces the auto-inhibitory domain (AID) from the active site of CaN, activating phosphatase activity. In the absence of calcium-loaded CaM, the RD is disordered, and binding of CaM induces folding in the RD. In order to provide mechanistic detail about the CaM–CaN interaction, we have undertaken an NMR study of the RD of CaN. Complete ¹³C, ¹⁵N and ¹H assignments of the RD of CaN were obtained using solution NMR spectroscopy. The backbone of RD has been assigned using a combination of ¹³C-detected CON-IPAP experiments as well as traditional HNCO, HNCA, HNCOCA and HNCACB-based 3D NMR spectroscopy. A ¹⁵N-resolved TOCSY experiment has been used to assign Hα and Hβ chemical shifts.

Original language	English
Pages (from-to)	215-219
Number of pages	5
Journal	Biomolecular NMR Assignments
Volume	11
Issue number	2
DOIs	https://doi.org/10.1007/s12104-017-9751-x
State	Published - Oct 1 2017

Bibliographical note

Funding Information:
We thank Scott Showalter and Jinfa Ying for assistance with CON spectral acquisition and data processing. We would like to thank Anthony Persechini (University of Missouri at Kansas City, Kansas City, MO) for the pETCaMI vector. Research reported in this publication was supported by the National Institutes of General Medical Sciences of the National Institutes of Health under Award No. R15GM113152. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

Publisher Copyright:
© 2017, Springer Science+Business Media B.V.

Keywords

Calcineurin
Calmodulin Binding
Intrinsically Disordered Protein

ASJC Scopus subject areas

Structural Biology
Biochemistry

Access to Document

10.1007/s12104-017-9751-x

Cite this

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title = "1H, 15N, and 13C chemical shift assignments of the regulatory domain of human calcineurin",

abstract = "Calcineurin (CaN) plays an important role in T-cell activation, cardiac system development and nervous system function. Previous studies have demonstrated that the regulatory domain (RD) of CaN binds calmodulin (CaM) towards the N-terminal end. Calcium-loaded CaM activates the serine/threonine phosphatase activity of CaN by binding to the RD, although the mechanistic details of this interaction remain unclear. It is thought that CaM binding at the RD displaces the auto-inhibitory domain (AID) from the active site of CaN, activating phosphatase activity. In the absence of calcium-loaded CaM, the RD is disordered, and binding of CaM induces folding in the RD. In order to provide mechanistic detail about the CaM–CaN interaction, we have undertaken an NMR study of the RD of CaN. Complete 13C, 15N and 1H assignments of the RD of CaN were obtained using solution NMR spectroscopy. The backbone of RD has been assigned using a combination of 13C-detected CON-IPAP experiments as well as traditional HNCO, HNCA, HNCOCA and HNCACB-based 3D NMR spectroscopy. A 15N-resolved TOCSY experiment has been used to assign Hα and Hβ chemical shifts.",

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note = "Funding Information: We thank Scott Showalter and Jinfa Ying for assistance with CON spectral acquisition and data processing. We would like to thank Anthony Persechini (University of Missouri at Kansas City, Kansas City, MO) for the pETCaMI vector. Research reported in this publication was supported by the National Institutes of General Medical Sciences of the National Institutes of Health under Award No. R15GM113152. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. Publisher Copyright: {\textcopyright} 2017, Springer Science+Business Media B.V.",

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AU - Tata, Sri Ramya

AU - Hunt, John

AU - Cook, Erik C.

AU - Creamer, Trevor P.

AU - Fitzkee, Nicholas C.

N1 - Funding Information: We thank Scott Showalter and Jinfa Ying for assistance with CON spectral acquisition and data processing. We would like to thank Anthony Persechini (University of Missouri at Kansas City, Kansas City, MO) for the pETCaMI vector. Research reported in this publication was supported by the National Institutes of General Medical Sciences of the National Institutes of Health under Award No. R15GM113152. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. Publisher Copyright: © 2017, Springer Science+Business Media B.V.

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N2 - Calcineurin (CaN) plays an important role in T-cell activation, cardiac system development and nervous system function. Previous studies have demonstrated that the regulatory domain (RD) of CaN binds calmodulin (CaM) towards the N-terminal end. Calcium-loaded CaM activates the serine/threonine phosphatase activity of CaN by binding to the RD, although the mechanistic details of this interaction remain unclear. It is thought that CaM binding at the RD displaces the auto-inhibitory domain (AID) from the active site of CaN, activating phosphatase activity. In the absence of calcium-loaded CaM, the RD is disordered, and binding of CaM induces folding in the RD. In order to provide mechanistic detail about the CaM–CaN interaction, we have undertaken an NMR study of the RD of CaN. Complete 13C, 15N and 1H assignments of the RD of CaN were obtained using solution NMR spectroscopy. The backbone of RD has been assigned using a combination of 13C-detected CON-IPAP experiments as well as traditional HNCO, HNCA, HNCOCA and HNCACB-based 3D NMR spectroscopy. A 15N-resolved TOCSY experiment has been used to assign Hα and Hβ chemical shifts.

AB - Calcineurin (CaN) plays an important role in T-cell activation, cardiac system development and nervous system function. Previous studies have demonstrated that the regulatory domain (RD) of CaN binds calmodulin (CaM) towards the N-terminal end. Calcium-loaded CaM activates the serine/threonine phosphatase activity of CaN by binding to the RD, although the mechanistic details of this interaction remain unclear. It is thought that CaM binding at the RD displaces the auto-inhibitory domain (AID) from the active site of CaN, activating phosphatase activity. In the absence of calcium-loaded CaM, the RD is disordered, and binding of CaM induces folding in the RD. In order to provide mechanistic detail about the CaM–CaN interaction, we have undertaken an NMR study of the RD of CaN. Complete 13C, 15N and 1H assignments of the RD of CaN were obtained using solution NMR spectroscopy. The backbone of RD has been assigned using a combination of 13C-detected CON-IPAP experiments as well as traditional HNCO, HNCA, HNCOCA and HNCACB-based 3D NMR spectroscopy. A 15N-resolved TOCSY experiment has been used to assign Hα and Hβ chemical shifts.

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