A Mechanism of Calmodulin Modulation of the Human Cardiac Sodium Channel

Christopher N. Johnson; Franck Potet; Matthew K. Thompson; Brett M. Kroncke; Andrew M. Glazer; Markus W. Voehler; Bjorn C. Knollmann; Alfred L. George; Walter J. Chazin

doi:10.1016/j.str.2018.03.005

A Mechanism of Calmodulin Modulation of the Human Cardiac Sodium Channel

Christopher N. Johnson^*, Franck Potet, Matthew K. Thompson, Brett M. Kroncke, Andrew M. Glazer, Markus W. Voehler, Bjorn C. Knollmann, Alfred L. George, Walter J. Chazin

^*Corresponding author for this work

Pharmacology

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

40 Scopus citations

Abstract

The function of the human cardiac sodium channel (Na_V1.5) is modulated by the Ca²⁺ sensor calmodulin (CaM), but the underlying mechanism(s) are controversial and poorly defined. CaM has been reported to bind in a Ca²⁺-dependent manner to two sites in the intracellular loop that is critical for inactivation of Na_V1.5 (inactivation gate [IG]). The affinity of CaM for the complete IG was significantly stronger than that of fragments that lacked both complete binding sites. Structural analysis by nuclear magnetic resonance, crystallographic, and scattering approaches revealed that CaM simultaneously engages both IG sites using an extended configuration. Patch-clamp recordings for wild-type and mutant channels with an impaired CaM-IG interaction revealed CaM binding to the IG promotes recovery from inactivation while impeding the kinetics of inactivation. Models of full-length Na_V1.5 suggest that CaM binding to the IG directly modulates channel function by destabilizing the inactivated state, which would promote resetting of the IG after channels close. The mechanism of calcium regulation of cardiac sodium channel (Na_V1.5) gating is much debated. Johnson et al. define a strong, calcium-dependent interaction between the intracellular calcium sensor calmodulin and the gate controlling inactivation of Na_V1.5. They find that calmodulin binding to the gate alters the recovery from inactivation.

Original language	English (US)
Pages (from-to)	683-694.e3
Journal	Structure
Volume	26
Issue number	5
DOIs	https://doi.org/10.1016/j.str.2018.03.005
State	Published - May 1 2018

Funding

We thank Greg Hura for assistance with processing SAXS data and Madeline Shea for providing plasmids of individual CaM domains. C.N.J. was supported by: NIH 2T32NS007491 , AHA postdoctoral fellowship 13POST14380036, and NIH 5 F32 HL117612 . This work was also supported in part by grants to A.L.G. (NIH R01 HL083374) and B.C.K. (NIH HL071670). Research on Ca 2+ binding proteins in the Chazin laboratory is supported by an endowed chair from Vanderbilt University and in part by NIH grant R35 GM118089. NMR instrumentation was supported by grants from the NSF (0922862), NIH (S10 RR025677), and Vanderbilt University matching funds. X-ray diffraction data were collected at the Life Sciences Collaborative Access Team beamline 21-ID-D at the Advanced Photon Source, Argonne National Laboratory operated for the DOE Office of Science by Argonne National Laboratory under contract no. DE-AC02-06CH11357. SAXS data were collected at the Advanced Light Source SIBYLS beamline, supported by DOE Office of Biological and Environmental Research and NIH (MINOS R01 GM105404; S10 OD018483). All software was utilized through the SBGRID consortium supported in part by NSF (MCB 0639193; EAGER 1448069). We thank Greg Hura for assistance with processing SAXS data and Madeline Shea for providing plasmids of individual CaM domains. C.N.J. was supported by: NIH 2T32NS007491, AHA postdoctoral fellowship 13POST14380036, and NIH 5 F32 HL117612. This work was also supported in part by grants to A.L.G. (NIH R01 HL083374) and B.C.K. (NIH HL071670). Research on Ca2+ binding proteins in the Chazin laboratory is supported by an endowed chair from Vanderbilt University and in part by NIH grant R35 GM118089. NMR instrumentation was supported by grants from the NSF (0922862), NIH (S10 RR025677), and Vanderbilt University matching funds. X-ray diffraction data were collected at the Life Sciences Collaborative Access Team beamline 21-ID-D at the Advanced Photon Source, Argonne National Laboratory operated for the DOE Office of Science by Argonne National Laboratory under contract no. DE-AC02-06CH11357. SAXS data were collected at the Advanced Light Source SIBYLS beamline, supported by DOE Office of Biological and Environmental Research and NIH (MINOS R01 GM105404; S10 OD018483). All software was utilized through the SBGRID consortium supported in part by NSF (MCB 0639193; EAGER 1448069).

Keywords

X-ray crystallography
calcium regulation
cardiac sodium channel (NaV1.5)
small angle X-ray scattering (SAXS)
solution NMR spectroscopy
structural biology
whole-cell patch-clamp electrophysiology

ASJC Scopus subject areas

Structural Biology
Molecular Biology

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10.1016/j.str.2018.03.005

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title = "A Mechanism of Calmodulin Modulation of the Human Cardiac Sodium Channel",

abstract = "The function of the human cardiac sodium channel (NaV1.5) is modulated by the Ca2+ sensor calmodulin (CaM), but the underlying mechanism(s) are controversial and poorly defined. CaM has been reported to bind in a Ca2+-dependent manner to two sites in the intracellular loop that is critical for inactivation of NaV1.5 (inactivation gate [IG]). The affinity of CaM for the complete IG was significantly stronger than that of fragments that lacked both complete binding sites. Structural analysis by nuclear magnetic resonance, crystallographic, and scattering approaches revealed that CaM simultaneously engages both IG sites using an extended configuration. Patch-clamp recordings for wild-type and mutant channels with an impaired CaM-IG interaction revealed CaM binding to the IG promotes recovery from inactivation while impeding the kinetics of inactivation. Models of full-length NaV1.5 suggest that CaM binding to the IG directly modulates channel function by destabilizing the inactivated state, which would promote resetting of the IG after channels close. The mechanism of calcium regulation of cardiac sodium channel (NaV1.5) gating is much debated. Johnson et al. define a strong, calcium-dependent interaction between the intracellular calcium sensor calmodulin and the gate controlling inactivation of NaV1.5. They find that calmodulin binding to the gate alters the recovery from inactivation.",

keywords = "X-ray crystallography, calcium regulation, cardiac sodium channel (NaV1.5), small angle X-ray scattering (SAXS), solution NMR spectroscopy, structural biology, whole-cell patch-clamp electrophysiology",

author = "Johnson, {Christopher N.} and Franck Potet and Thompson, {Matthew K.} and Kroncke, {Brett M.} and Glazer, {Andrew M.} and Voehler, {Markus W.} and Knollmann, {Bjorn C.} and George, {Alfred L.} and Chazin, {Walter J.}",

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AU - Glazer, Andrew M.

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AB - The function of the human cardiac sodium channel (NaV1.5) is modulated by the Ca2+ sensor calmodulin (CaM), but the underlying mechanism(s) are controversial and poorly defined. CaM has been reported to bind in a Ca2+-dependent manner to two sites in the intracellular loop that is critical for inactivation of NaV1.5 (inactivation gate [IG]). The affinity of CaM for the complete IG was significantly stronger than that of fragments that lacked both complete binding sites. Structural analysis by nuclear magnetic resonance, crystallographic, and scattering approaches revealed that CaM simultaneously engages both IG sites using an extended configuration. Patch-clamp recordings for wild-type and mutant channels with an impaired CaM-IG interaction revealed CaM binding to the IG promotes recovery from inactivation while impeding the kinetics of inactivation. Models of full-length NaV1.5 suggest that CaM binding to the IG directly modulates channel function by destabilizing the inactivated state, which would promote resetting of the IG after channels close. The mechanism of calcium regulation of cardiac sodium channel (NaV1.5) gating is much debated. Johnson et al. define a strong, calcium-dependent interaction between the intracellular calcium sensor calmodulin and the gate controlling inactivation of NaV1.5. They find that calmodulin binding to the gate alters the recovery from inactivation.

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A Mechanism of Calmodulin Modulation of the Human Cardiac Sodium Channel

Abstract

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