NaV1.2 EFL domain allosterically enhances Ca2+ binding to sites I and II of WT and pathogenic calmodulin mutants bound to the channel CTD

Ryan Mahling; Liam Hovey; Holly M Isbell; Dagan C Marx; Mark S Miller; Adina M Kilpatrick; Lisa D Weaver; Jesse B Yoder; Elaine H Kim; Corinne N.J Andresen; Shuxiang Li; Madeline A Shea

doi:10.1016/j.str.2021.03.002

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NaV1.2 EFL domain allosterically enhances Ca2+ binding to sites I and II of WT and pathogenic calmodulin mutants bound to the channel CTD

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NaV1.2 EFL domain allosterically enhances Ca2+ binding to sites I and II of WT and pathogenic calmodulin mutants bound to the channel CTD

Ryan Mahling, Liam Hovey, Holly M Isbell, Dagan C Marx, Mark S Miller, Adina M Kilpatrick, Lisa D Weaver, Jesse B Yoder, Elaine H Kim, Corinne N.J Andresen, …

Structure (London), Vol.29(12), pp.1339-1356.e7

03/2021

DOI: 10.1016/j.str.2021.03.002

PMCID: PMC8458505

PMID: 33770503

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Abstract

Neuronal voltage-gated sodium channel NaV1.2 C-terminal domain (CTD) binds calmodulin (CaM) constitutively at its IQ motif. A solution structure (6BUT) and other NMR evidence showed that the CaM N domain (CaMN) is structurally independent of the C-domain (CaMC) whether CaM is bound to the NaV1.2IQp (1,901–1,927) or NaV1.2CTD (1,777–1,937) with or without calcium. However, in the CaM + NaV1.2CTD complex, the Ca2+ affinity of CaMN was more favorable than in free CaM, while Ca2+ affinity for CaMC was weaker than in the CaM + NaV1.2IQp complex. The CTD EF-like (EFL) domain allosterically widened the energetic gap between CaM domains. Cardiomyopathy-associated CaM mutants (N53I(N54I), D95V(D96V), A102V(A103V), E104A(E105A), D129G(D130G), and F141L(F142L)) all bound the NaV1.2 IQ motif favorably under resting (apo) conditions and bound calcium normally at CaMN sites. However, only N53I and A102V bound calcium at CaMC sites at [Ca2+] < 100 μM. Thus, they are expected to respond like wild-type CaM to Ca2+ spikes in excitable cells. [Display omitted] •In solution, only the IQ motif of NaV1.2 CTD(1,777–1,937) contacts CaM ± Ca2+•Ca2+ changes interfaces of CaM + NaV1.2-IQp and CaM + NaV1.2-CTD complexes identically•Ca2+ binds to N-domain sites of CaM + NaV1.2-CTD preferentially relative to free CaM•Apo pathogenic CaM mutants bind the NaV1.2 CTD as well or better than WT CaM The NaV1.2 C-terminal domain (CTD) constitutively binds the calmodulin C-domain (CaMC). Mahling et al. use NMR and thermodynamics to show the CTD's EF-like domain improves Ca2+ binding to WT calmodulin N domain (CaMN), while Ca2+-binding rotates CaMC. Pathogenic CaMC mutants resist rotation, but their CaMN sites behave like WT.

Molecular Recognition

NMR

biosensor

titration

free energy

binding

FRET

linkage

voltage-gated sodium channel

allostery

affinity

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Title: Subtitle: NaV1.2 EFL domain allosterically enhances Ca2+ binding to sites I and II of WT and pathogenic calmodulin mutants bound to the channel CTD
Creators: Ryan Mahling - Department of Biochemistry, Carver College of Medicine, University of Iowa, Iowa City, IA 52242-1109, USA
Liam Hovey - Department of Biochemistry, Carver College of Medicine, University of Iowa, Iowa City, IA 52242-1109, USA
Holly M Isbell - Department of Biochemistry, Carver College of Medicine, University of Iowa, Iowa City, IA 52242-1109, USA
Dagan C Marx - Department of Biochemistry, Carver College of Medicine, University of Iowa, Iowa City, IA 52242-1109, USA
Mark S Miller - Department of Biochemistry, Carver College of Medicine, University of Iowa, Iowa City, IA 52242-1109, USA
Adina M Kilpatrick - Department of Physics and Astronomy, Drake University, Des Moines, IA 50311-4516, USA
Lisa D Weaver - Department of Biochemistry, Carver College of Medicine, University of Iowa, Iowa City, IA 52242-1109, USA
Jesse B Yoder - Department of Biochemistry, Carver College of Medicine, University of Iowa, Iowa City, IA 52242-1109, USA
Elaine H Kim - Department of Biochemistry, Carver College of Medicine, University of Iowa, Iowa City, IA 52242-1109, USA
Corinne N.J Andresen - Department of Biochemistry, Carver College of Medicine, University of Iowa, Iowa City, IA 52242-1109, USA
Shuxiang Li - Department of Biochemistry, Carver College of Medicine, University of Iowa, Iowa City, IA 52242-1109, USA
Madeline A Shea - Department of Biochemistry, Carver College of Medicine, University of Iowa, Iowa City, IA 52242-1109, USA
Resource Type: Journal article
Publication Details: Structure (London), Vol.29(12), pp.1339-1356.e7
DOI: 10.1016/j.str.2021.03.002
PMID: 33770503
PMCID: PMC8458505
NLM abbreviation: Structure
ISSN: 0969-2126
eISSN: 1878-4186
Publisher: Elsevier Ltd
Grant note: DOI: 10.13039/100008893, name: University of Iowa; DOI: 10.13039/100015515, name: University of Iowa Roy J and Lucille A Carver College of Medicine; DOI: 10.13039/100000002, name: NIH
Language: English
Date published: 03/2021
Academic Unit: Molecular Physiology and Biophysics; Iowa Neuroscience Institute; Biochemistry and Molecular Biology
Record Identifier: 9984070456602771

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